The Importance Of Animal Uses In Animals

The Importance Of Animal Uses In Animals
Humans have used animals for food, transport, companionship and experimental research to develop
medicine and investigate the structure and function of the body. This dates back to 130 AD, where
the commonly used animals were pigs, monkeys, and dogs (Vaumans, 2005). Dogs are the species
most frequently used in the animal models and are referred to as "default non–rodent" (Smith et al.,
2002). Especially, in toxicology studies, dogs are the preferred choice due to their similarities of
human toxicity (Smith et al., 2002). Today, 75–100 million vertebrates are used per year worldwide
for research, teaching, and testing (Vaumans, 2005). In many countries it is mandatory to grade the
level of discomfort and pain for the laboratory animal. Animals suffering from discomfort and/or
stress can have a change in their endocrinology and immunology results (Vaumans, 2005). By
testing oxytocin concentrations in dogs can help facilitate the transition into new social settings and
into new laboratory settings (Stallings, 2015). In addition, oxytocin plays a role in non–social
observations such as learning, anxiety, feeding, and pain (Stallings, 2015). All laboratory animals
fall under the Animal Welfare Act, which promotes animal welfare of research and laboratory
animals and promotes anti–cruelty laws. Currently, according to American Association for
Laboratory Animal Science, (AALAS) animal welfare and assess laboratory animals' pain and
distress is conducted by a trained laboratory
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Lab Report On Luciferase
Introduction
In this experiment, the enzyme Luciferase (Luc) was purified using differential centrifugation,
precipitation, and a simplified chromatography procedure, followed by an enzyme activity assay
using the enzyme's natural substrate, Luciferin, to confirm the presence of Luc.
Luciferase is a 61 kD enzyme naturally found in the tails of fireflies, a member of the Lampyridae
family. Luc acts as a catalyst in an oxidation–reduction reaction with Luciferin, which allows these
insects to exhibit bioluminescence for mating and communication ("Firefly Luciferase," n.d.). In this
reaction, Luciferin is adenylated and oxidized to produce Oxyluciferin, along with carbon dioxide,
adenosine monophosphate, and light ("Luciferase Mechanism," ... Show more content on
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Following the purification process, the enzyme activity assay should reveal luminescence in the
supernatant fraction of the tail sample. SDS–PAGE of the samples should reveal a band at 61 kD in
the tail sample and no distinct banding in the head sample.
Materials and Methods
Protein Purification:
To begin, samples of both firefly heads and firefly tails were divided amongst the groups. Our group
was assigned the firefly head to use as a negative control, while the other was assigned to the tail.
The firefly tissue was lysed by grinding with a pestle in the tube (Tube A) both before and after
adding 250 L of Lysis buffer (1X PBS with 1% SDS). This mixture was then sonicated for two
minutes to complete the lysing process.
The tubes were then centrifuged at 1,000 rpm for 3 minutes to remove the large tissue pieces. The
supernatant of this tube contained the soluble compounds and proteins that were present in the
cytosol of the cells present in the tissue. The supernatant was removed using a micropipette and
transferred to a new 1.5 mL–Eppendorf tube (Tube B), and the pellet Tube A was discarded.
The supernatant in Tube B was then mixed with 1 mL of ice cold acetone in order to separate the
proteins from the other compounds present in the aqueous solution. Tube B was spun in the
centrifuge at 10,000 rpm for 2 minutes. The resulting supernatant was removed and discarded to
isolate the pellet, containing the

Questions On Materials And Methods
2– Materials and methods 1. Snails B. alexandrina snails (8 – 10 mm) from Medical Malacology
Laboratory, Theodor Bilharz Research Institute (TBRI), Giza, Egypt were used. Pieces of
polyethylene sheets were put into the aquaria to collect egg masses. 2. Synthetic insecticide: Match
5% EC: Lufenuron 5% EC (Registration No., 609) was purchased from Syngenta (AGRO–Egypt).
3. Bioassay tests: 3.1. Molluscicidal screening: A stock solution of 1000 ppm was prepared from the
insecticide on the basis of W/V using dechlorinated tap water. To calculate LC50 and LC90, Series
of concentrations were prepared and ten B. alexandrina snails were placed in beakers for each
concentration (WHO, 1983) and another group in dechlorinated water as control. Three replicates
were prepared for each group and control, then, snails' mortality was recorded. 3.1.1. Collection of
hemolymph: To collect the hemolymph, a small portion of the shell which situated directly above
the heart of snails was removed and a capillary tube was inserted into the heart (Nduka and
Harrison, 1980). a– Total hemocytes count: The number of cells was counted using a Bürker– Turk
hemocytometer (Van der knap et al., 1981) by using 10 µl of hemolymph of each group. b–
Differential hemocytes count: Hemocytes monolayers were prepared according to the method of
Abdul–Salam & Michelson (1980). c– Biochemical Assays: Activities of aspartate aminotransferase
(AST) and alanine aminotransferase (ALT) in

A Diagnostic Tool For Renal Disease And Hypertensive...
Introduction Proteinuria is often used to detect the presence of any type of proteins in the urine and
serves as a diagnostic tool for renal disease and hypertensive disorders. Among those disorders, pre–
eclampsia is characterized by low renal perfusion and leakage of proteins in urine due to a damaged
glomerular base membrane (1). This condition is associated with a high risk of maternal
complications and prenatal mortality. Pre–eclampsia is usually defined as a combination of
hypertension, edema and proteinuria after 20 weeks of gestation and is determined by a threshold of
300mg/24 hours of proteins (2). Normal urine excretion is up to 150mg/day (3). The most common
test used in antenatal care to screen proteinuria is the visual reagent dipstick. It is a semi–
quantitative and colorimetric method. The stick changes colour when the reagent
(tetrabromophenol) binds to the proteins. The binding is pH dependent (albumin binds at a pH
between 5 and 7, other proteins bind only at a pH below). If the urinalysis by the dipstick is positive,
a 24 hour–urine collection is performed in order to confirm the proteinuria and to estimate the total
amount of protein (4). Another method available for detecting proteinuria is called the Bradford
assay. This quantitative method is based on the binding of proteins with the dye Coomassie Blue
which increases the absorption of the dye at 595nm. The absorbance measured by
spectrophotometry is related to the amount of proteins present in the

Biology Case Study
Materials and methods
The levels of microRNA–22–3p (miR–22–3p) and TP63 were analyzed by RT–qPCR and/or
Western blot. Cell proliferation was measured using MTT assay. Cell invasion and migration were
evaluated using Transwell assay chambers. Prediction of a regulatory relationship between
microRNA–22–3p and 3'–UTR of TP63 mRNA was performed by a bioinformatics algorithm and
confirmed by a dual luciferase reporter assay.
OC cell lines
Lung cancer cell lines H292, PC–9, CL1–5, A549 and normal NHBE cells were used in this study.
All of Lung cancer and normaL ceLL Lines – were provided by ATCC. H292, PC–9, CL1–5, A549
and NHBE cells were cultured in RPMI–1640 medium supplemented with 10% fetal bovine serum
(FBS).
MiRNA and RNA interference ... Show more content on Helpwriting.net ...
The experiments were repeated thrice.
Cell migration and invasion assays
In vitro cell migration and invasion were examined using the Boyden Chamber Assays. For the cell
migration assay, 30000 cells in 100 μl DMEM medium devoid of fetal bovine serum (FBS) and a
given concentration of cetuximab were cultured on a polycarbonate membrane insert coated with
fibronectin in a Boyden Chamber (Essen BioScience, UK). In the lower chamber, 500 μl DMEM
with 10% FBS was added as chemoattractant. After the cells were incubated for 6 h at 37 °C in a 5%
CO2 atmosphere, the insert was rinsed using PBS and cells on the upper side of the insert were
gently removed using a cotton swab. Subsequently, cells that adhered to the lower side were fixed
with methanol, stained with crystal violet solution and counted under a microscope in five
predetermined fields (×200). All assays were independently repeated at least thrice. The same
protocol as described above was used for the cell invasion assay, except that the transwell filter
membranes were pre–coated with 24 μg/μl matrigel (R&D Systems, Inc., Minneapolis, MN, USA)
and the cells were incubated for 8 h at 37 °C in a 5% CO2 atmosphere.
RNA isolation, reverse transcription and RT–qPCR
RNA was extracted from the OC cells using the Trizol reagent. For miR–22–3p, RNA was
transcribed into complementary DNA and amplified using miRNA PrimeScript RT Enzyme Mix kit
according to the manufacturer's instructions

Advantages And Disadvantages Of Reverse Sequence Syphilis...
Introduction
The traditional algorithm for syphilis screening has involved use of a manual non–treponemal test
(e.g. rapid plasma reagin, RPR) followed by confirmation with a treponemal test (e.g. Treponema
pallidum particle agglutination, TP–PA). Non–treponemal tests are inexpensive, but require
significant hands–on time by laboratory personnel and produce subjective results. High volume
laboratories are increasingly using a reverse sequence algorithm, using partially or fully automated
treponemal tests (e.g. enzyme immunoassay–EIA, chemiluminescene immunoassay–CIA, multibead
immunoassay–MBIA), followed by a non–treponemal test (e.g. RPR, VDRL) if reactive (1).
Potential advantages of the reverse sequence algorithm include reduced labor ... Show more content
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Our objective was to determine signal strength cutoffs associated with 98%, 99%, and 100%
reactive confirmatory testing (TP–PA or fluorescent treponemal antibody absorbed test, FTA–ABS).
For the four automated immunoassays we also attempted to establish a signal strength cutoff
associated with a consensus of at least 4/7 reactive treponemal tests (the index test plus at least 3
others). The findings from this study will inform laboratory guidelines on use of confirmatory
treponemal testing for patients with initially discordant serology.
Results
Among 1995 specimens tested, 907 (45%) were categorized as true positives and 1087(54%) as true
negatives using a consensus of the test panel as the gold standard (see Materials and Methods for
case definitions). Table 1 includes results for positive agreement, negative agreement and Kappa's
statistics for the seven assays. Positive agreement was lowest for FTA–ABS (86.1%, 83.7–88.2%)
and highest for Centaur EIA (99.7%, 99.0–99.9%). Positive agreement

Cell Viaability Lab Report
experiment. (5%) HEPG2 R²=0.8354 PANC–1 R²= 0.9734 PC–3 R²= 0.9482 For each value shown
we can determine that there is a strong correlation between drug concentration and % cell viability.
This can tell us that as the drug concentration increases the % cell viability decreases. This shows
that the chemotherapeutic agent is having the desired effect. 4. What conclusions can you draw from
the results of the experiment? (10%) I can draw from these results that as the chemotherapeutic drug
concentration is increased the cell viability decreases. In HEPG2 the viability at the control was
100% but when the drug was added at concentration of 40µm the cell viability decreased to 2.62%.
From these examples we can deduce that the chemotherapeutic drug added is successful in its use
and that it is targeting the cancerous cells. The drug damages the genes present inside the cell line ...
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It achieves this my measuring the mitochondrial enzyme activity and as such is used to determine
the cytotoxicity of potential medical drugs. It measures the reduction of yellow 3–(4,5–
dimethythiazol–2–yl)–2,5–diphenyltetrazolium bromide (MTT) by mitochondrial succinate
dehydrogenase. The MTT starts as yellow but as it enters the cells and passes into the mitochondria
where it is reduced and changes in colour to a dark purple formazan product. The cells are then
solubilised with an organic solvent and released, solubilised formazan reagent is measured using a
spectrometer (Cree, 2011). Some errors that may occur while using this assay are that uneven
evaporation of culture fluid in the wells may cause erroneous results. The MTT solution is stable for
6 months when stored at 0°C but at 2–8°C for more than 2 weeks may cause decomposition and
erroneous results may occur. Microbial contamination will contribute to cleavage of MTT and the
formation of MTT formazan, causing erroneous results (Sigma Aldrich,

Potential Therapeutics For Treating Type 2 Diabetes Mellitus
Bromophenols have been identified as potential therapeutics for treating type 2 diabetes mellitus
(T2DM) due to their suspected tyrosine phosphatase 1B (PTP1B) inhibitory activity. A new series of
bromophenol analogues will be synthesised and their PTP1B inhibitory activity will be tested in in
vitro enzymatic assays to elucidate their mechanism of action as PTP1B inhibitors.
Background and Introduction
Type 2 diabetes mellitus (T2DM) is a metabolic disease that is caused by insulin resistance of the
peripheral tissues and impaired insulin secretion of the pancreatic β cells.1 Both conditions result in
reduced glycemic control, which can lead to hyperglycemia. Despite the availability of a wide range
of anti–diabetic drugs to treat T2DM, ... Show more content on Helpwriting.net ...
Bromophenols have attracted much attention as potential PTP1B inhibitors due to their suspected
anti–diabetic activity.4 Bromophenols occur naturally in marine organisms such as algae, mollusks
and jellyfish. (±)–Polysiphenol (1, figure 1) is an example of a bromophenol that can be extracted
from the marine algae Polisiphonia Ferulecea.5 After the Braddock group successfully achieved the
first total synthesis of (±)–polysiphenol and the separation of its enantiomers, they collaborated with
the Woscholski group to test their inhibitory activity in PTP1B enzymatic assays.6,7 Both
enantiomers of polysiphenol were essentially found to be identical in PTP1B inhibitory activity.
Thus polysiphenol, as single enantiomers, cannot be responsible for the observed inhibition. This
implicates a chemical change of polysiphenol into a non–chiral form is likely to take place under the
conditions of the assay. One possibility is the formation of a para–quinone methide species (4, figure
1), which in turn can tautomerise into an ortho–quinone methide species (5, figure 1). Thus there are
various possible reactive species that might be responsible for the observed inhibition of PTP1B.
The formation of quinones highlights a red flag in drug discovery due to their highly reactive nature.
Quinones are Michael acceptors that are known to cause toxicity in vivo by covalently binding to
nucleophilic sites on proteins and DNA.8 Consequently, quinones

DAPI Stain Living Cells
Introduction
The number of cells is closely regulated by the rate of cell division and the rate of cell death. When
cells become unneeded, they activate an intracellular death program and commit suicide. This
process is often known as apoptosis. Cells die for numerous reasons. For instance, cell death
regulates cell numbers and can adjust the number of cells needed for a particular function. When a
structure a cell forms is no longer needed, the cell will die. Cells can become damaged or stressed
and trigger an activation from within the cell to kill itself. Proteolytic enzymes called capases
mediate apoptosis by triggering cell death by cleaving proteins in the cytoplasm and nucleus. The
triggering process is initiated by death signals by either extracellular or intracellular signals.
Response to signals coming from ... Show more content on Helpwriting.net ...
Specific protein location and morphology of organelles can be found using DAPI stains. The DAPI
stain is classified as a nuclear stain that binds A–T regions of DNA. DAPI significantly increases the
fluorescent properties and can be identified as a blue fluorescence of DAPI when processed by UV
light. The only implications include using fixed cells. Using a Mitotracker Red CMXRos, you can
stain living cells mitochondria. Requiring an active mitochondrial membrane potential the
mitochondria oxidizes the Mitotracker probe. Fluorescence property is maintained after fixation. An
ER tracker viable stain uses the bodipy fluorophore. The glibenclamide probe selectively binds
sulphonylurea receptors of ATP sensitive K+ channels that are localized in the ER. Another viable
stain used was the Lysotracker. Its probes are very selective for acidic compartments. Its probe is
retained in the lysosome via protonation of the probe. These different stains are identified as the
mitotracker will be red, lysotracker is green and the Hoescht stain will be

Resistance Of The Mutant Oncogenes
Introduction
One in five deaths today is a result of some form of cancer. This terrible disease stems from
mutations in body cells, which interfere with the cells' normal regulation of growth, division, and
death. Cells can make occasional mistakes and divide too early or not destroy themselves at the right
time (Alberts et al. 2014). The Hayflick limit, described in 1961 by Leonard Hayflick, states that
normal human cells can divide for about 50 times (Watts 2011). In cancers, though, this limit does
not exist, and cells continually divide. Occasional mistakes do not have tremendous consequences
on large, multicellular organisms, however. When many cells begin to make these mistakes, though,
tumorigenesis and cancer can occur. ... Show more content on Helpwriting.net ...
The pigment protects skin cells, particularly the DNA of the cells, from damaging ultraviolet
radiation from the sun, and has been shown to reduce the frequency of radiation–induced mutations
and to completely remove low–dose radiation effects (Mosse et al. 2000). Within melanocytes are
melanosomes, organelles in which melanin production occurs. The melanosomes have 4 stages of
maturity, starting with stage 1, or premelanosomes, and progressing to stage 4, or mature
melanosomes. The stages can be determined based on the composition of the vesicles in the cells
(Setaluri 2003). The mature melanosomes then branch off from the dendritic tips of the melanocytes
and travel to keratinocytes, keratin–producing cells (Seiberg 2001).
The chemotherapy drug methotrexate is an alkaloid that has been used as a treatment for acute
lymphocytic leukemia, non–Hodgkin's lymphoma, osteosarcoma, choriocarcinoma, head and neck
cancer, and breast cancer. Methotrexate is an antimetabolite that acts as a folic acid antagonist.
Components of folate vitamins are required for DNA synthesis and cell replication, so by inhibiting
folic acid and other folates, methotrexate slows the DNA synthesis and division of certain cancer
cells, and impacts cell proliferation (Jolviet et al. 1983). The estimated half maximal effective
concentration (EC50) for methotrexate was found to be 0.00339μM for human breast cancer MDA–
MB–231

LDH Purification lab Report
PURIFICATION OF LACTATE DEHYDROGENASE FROM CHICKEN MUSCLE TISSUE
Abstract
The enzyme lactate dehydrogenase (LDH) catalyzes the last step of anaerobic glycolysis that is
important for the normal function of the body. Purification of LDH is essential to understand its
structure and function. The purpose of this experiment was to extract and purify LDH enzyme from
chicken muscle tissue using a variety of various. Analytical methods such as activity and protein
assay were employed to determine the presence and purity of LDH. The cells were initially
disrupted and proteins were solubilized. LDH was purified from the ammonium sulfate precipitated
protein mixture by affinity chromatography and its activity was studied by ... Show more content on
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10 microliters of the sample is then added and the assay absorption is measured at 340nm. If
absorbance was above 1.5, samples were diluted. Protein Assay: The Pierce BCA Protein Assay
(Thermo Scientific) is a detergent–compatible formulation based on bicinchoninic acid (BCA) for
the colorimetric detection and quantitation of total protein concentration. A series of standard
solution of Bovine Serum Albumin (BSA) ranging from 0–2000 µg/ml was prepared from a stock
solution of 2 mg/ml BSA. 25ul of diluted crude (1:500, 1:250), desalted (1:100, 1:50), and 6 peak
fractions from cibarcon blue column (1:10, 1:5) were loaded in microplate along with 175ul of BCA
working reagent. Microplate was incubated for 30min at 370C and then the absorbance was
measured at 562nm.
Results/Discussion The purpose of this experiment was to extract and purify LDH enzyme from
chicken muscle tissue using a variety of techniques including homogenization, ammonium sulfate
precipitation, dialysis, and affinity chromatography. Activity and Protein assay were used to track
the overall amount of LDH present in the samples. Crude Extraction: Chicken muscle tissue was
homogenized in a blender with cold extraction buffer in order to lyse cells, releasing LDH into

Use Of Small Volume Fluids On Scale Of The Field Of...
ABSTRACT
Microfluidics biochips are revolutionary devices in the field of clinical diagnostics, DNA analysis
and molecular biology. Biochips involve various fields of science and engineering i.e., physics,
chemistry, biochemistry, nanotechnology, fabrication technology and computer science. It uses small
volume fluids on scale of micro to nano liter for automatically carrying out the reactions needed for
some biochemical assays. In this report I have covered basic overview of biochips, kinds of
biochips, areas of research, existing problems and finally presented work done till now in area of
sample preparation specifically dilution and mixing Algorithms for Digital Microfluidics Biochips.
INTRODUCTION
Microfluidics biochips are emerging devices for analysis of biological assay, and have its potential
application in the field of healthcare because of its point–of–care diagnostic capability. These chips
are of very small size typically of few centimeters [4]. These chips also have its application in
Biological computing it can be used as sample preparation primitive which reduces the operations
time. DNA hybridization can be reduced to 4min from 24hrs [2] by using it. Due to its small size
and low cost it feasible to use in place of bulky convention diagnosis devices.
These biochips also used to monitor toxicity of air and water. It has many advantages such as micro
liter fluids amount required for assay operation, high speed analysis, less cost in

Purification of lactate Dehydrogenase
Purification of Lactate Dehydrogenase
Results and Discussion
Monitoring Assays Enzyme activity assay and Bradford dye binding protein assay was used.
Ammonium Sulfate Fractionation In order to isolate and purify Lactate Dehydrogenase (LDH), we
first extracted LDH from bovine muscle tissue. We minced 8 grams of meat and homogenized using
blender. The homogenate was centrifuged and the pellet consisting of membranes, organelles,
cytoskeletal components and structural fragments was discarded. The crude extract was subjected to
40% (NH4)2SO4 fractionation. The purpose of this is to remove molecules of lesser solubility than
LDH like lipids, fats and low soluble proteins. For this, 9.24 g of was slowly added to crude extract
while stirring reaching final solution saturation of 40%. The mixture was then centrifuged and the
pellet consisting the contaminants was discarded. From table I, the data indicate that the yield for
this was 88% with purification factor of 1.5. The total protein in 40% supernatant was 121 mg
which decreased from 210 mg present in crude extract while retaining 88% of LDH suggesting
some contaminant proteins were removed. Due to some discrepancies in protein data for crude
extract and 40% supernatant, we re–assayed these. Even though week 3 data may not be very
reliable, the crude extract assay from week 2 and 40% supernatant assay from week 3 gave the most
appropriate data with least error. The 40% supernatant was then subjected to 60%

Enzyme Catalysis And Enzyme Inhibition Essay
Enzyme catalysis and enzyme inhibition are two essential biological mechanisms of organisms. In
this experiment, WT–AP and MBP–AP enzyme are reacted with different concentrations of PNPP
substrate in SpectrovVis time based assays. From the change in absorbance over time data, and the
rates of the reactions are calculated, followed by the determination of the kinetic constants. Then,
the MBP–AP assays are repeated with two different concentrations of phosphate inhibitor and the
kinetic constants of the uninhibited enzymes are compared to those of the inhibited enzymes.
Results of the experiment show that the uninhibited WT–AP and MBP–AP enzymes yielded
expected kinetic constants according to a reference. However, the inhibited enzymes produced
kinetics constants that did not resemble a competitive inhibition, which was expected. Instead, the
inhibitor is observed to be a mixed inhibitor.
Introduction Enzymes are biological catalysts. Without them, biological reactions will not proceed
in a reasonable rate. For example, without a series of digestive enzymes, it can take 20 years to
digest a hamburger. Enzyme–catalyzed reactions proceed through an ES complex, in which the
substrate binds to the enzyme, gets transformed into the product, and releases into the environment.
For a specific enzyme, only one or a few different substrate molecules can bind in the proper
manner and produce a functional ES complex. The substrate must have a size, shape, and polarity
compatible with

The Spectrophotometer Determination of Protein...
The Spectrophotometer Determination Of Protein Concentrations And The Effects Sodium Dodecyl
Sulphate And Triton X–100 Have On Protein Concentration.
INTRODUCTION
Spectroscopy is used as a collective term for all the analytical techniques based on the interaction of
light and matter. Spectrophotometry is one of the branches of spectroscopy where we measure the
absorption of light by molecules that are in a gas or vapour state or dissolved molecules/ions
(Tombs, et.al, 1959). Spectroscopy is the use of electromagnetic radiation by a sample in order to
identify components of the sample (qualitative analysis) or to measure the amount of material in that
sample (quantitative analysis) (Tombs, et.al, 1959). Absorption is a process in which ... Show more
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MATERIALS AND METHODS
The BIORAD dye was firstly prepared by dissolving 0.225g of Coomassie Brilliant Blue G–250
(Merck, South Africa), in 67.5ml of Methanol (Riedel–de–Haёn, Germany) and 291ml of 85% (w/v)
Phosphoric Acid, diluted to 450ml with dH2O. Six test tubes were labelled SC1–6 and the 0.1mg/ml
stock of Bovine Serum Albumin (Roche Diagnostics, Germany) and dH2O was pipette into their
respective test tubes according to Table 1.
Table 1: Table of the BSA and dH2O volumes to be pipette and placed in the spectrophotometer to
obtain absorbencies for the standard curve
Test Tube Number Total Volume (μl) Protein (BSA) Stock (μl) H2O (μl) Amount of Protein (μg)
1 2400 0 2400 0
2 2400 50 2350 5
3 2400 100 2300 10
4 2400 150 2250 15
5 2400 200 2200 20
6 2400 250 2150 25
600μl of the BIORAD dye agent was pipette into each test tube making up the volume to 3000μl.
All the test tubes were then covered with Parafilm and placed in the Vortex (Lab Pro cc) for a few
seconds and they were incubated at room temperature for 5–60 minutes. Simultaneously another set
of test tubes were labelled X1–8 and pipette with 0.1mg/ml Lysozyme stock (Merck, Germany),
BSA, 1% (w/v) Sodium Dodecyl Sulphate prepared with dH2O, 1% (v/v)

Disadvantages Of Refractometer
INTRODUCTION
A refractometer is aneoptical device that takes advantage of the fact that light passing through a
liquid bends or refracts. The refractometer is a tool. It is used to help you select higher quality of
foods containing more vitamins and minerals. They are also used to determine the concentration of
dissolved solids in a solution. Cooking oil, molasses, syrup, and other thick liquids require a
refractometerqcalibrated to read 30–90 Brix. Refractometer also is used in wine making. (Harrill,
1998) refractometer is also used to test to quality of fruits and vegetables.
A hydrometer is an instrument used to measure the density of a liquid as compared to that of water.
Hydrometers usually consist of a calibrated glass tube ending in ... Show more content on
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Why?
Temperature is an essential parameter because the index of refraction is affected by temperature.
The speed of light in a liquid will increase when the temperature increases. For instance, according
to Hanson (2006), when there is 1°C increasing in temperature, many organic liquids the index of
refraction decreases by nearly %0.05
What are the advantages and disadvantages of both instruments in measuring total soluble solid
content? Refractometer advantages: İnexpensive instrument
Refractometer disadvantages: Tedious calibration No instrument protocol Operator dependent
readings( limited accuracy) Long and difficult thermosetting (water bath is required after each
experiment)
Hydrometer advantages: Quick instrument Simple method Inexpensive instrument
Hydrometer disadvantages; Breakable glassware especially be careful when it is dropped into
solution Difficult to clean and dry it needs to be sure that it is cleaned well because not enough to
clean can cause measurement mistake Operator dependent readings Large sample volume required
("Comparision of different measuring techniques for density and

The shape and magnitude of the UV spectra depends on the...
The shape and magnitude of the UV spectra depends on the composition of amino acid in each
protein. Due to the aromatic amino acid residues in the protein, the observed UV absorbance was
mainly in the 240 nm to 340 nm region. In Figure 1 to 3, the maximal absorbance of each protein
was approximately at 280 nm. The difference in magnitudes of the peak observed was linked to the
differences in the amino acid contents in each of the proteins. The peak of lysozyme was greater
than those of BSA and gelatin, because lysozyme has a greater number of tryptophan residues.
Lysozyme has six tryptophan residues, whereas BSA and gelatin has two and zero, respectively
(Department of Chemistry, 2014). Lysozyme has three times more tryptophan residues ... Show
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In order to examine the variation in sensitivity of lysozyme, gelatin, and BSA, the absorbance
reading was obtained using the Bradford and BCA assay
Gelatin is observed to have a lower absorbance reading than lysozyme in only the Bradford assay,
while in the BCA assay gelatin was observed to have about the same absorbance as lysozyme. In
Bradford assay, the color yield for proteins with higher content of tryptophan, tyrosine, or cysteine
residue, will be higher. The observed protein sensitivities did not correlates with the amino acid
content in the Bradford assay. As presented in Table I, the mole fractions of lysine and arginine in
BSA were 10 mole percent and 4 mole percent, respectively. The sensitivity should have been higher
in BSA than in the Bradford assy. However, the observed sensitivities did correlates with the amino
acid contents in the BCA assay. Bradford assay would be ideal if an accurate estimation of a newly
isolated protein using BSA as a standard. With the data obtain from the Bradford assay, it shows that
it was more sensitive and quicker, also it had less interference from substance that were possibly
present in the protein solution, expect for the interfering substance SDS.
The interference of RNA and ME in the Bradford assay was compensated by using the blank,
because the difference between the addition of the protein and the non–protein blank

The Effect Of Alkaline Phosphatase On Para...
A SPECTROPHOTOMETRIC ASSAY OF THE ACTIVITY OF ALKALINE PHOSPHATASE ON
PARA–NITROPHENYLPHOSPHATE IN THE PRODUCTION OF PARA–NITROPHENOL
M. Smit1
1 Department of Biological Sciences, University of Cape Town, Rondebosch
Introduction:
Alkaline phosphatase is a biologically important enzyme, catalysing the hydrolysis of phosphate
groups in organic compounds through the process of dephosphorylation. para–Nitrophenylphosphate
(pPNP) is an example of a chromogenic substrate of this enzyme, because the product of hydrolysis
of pPNP is the yellow para–nitrophenol (pNP) compound along with inorganic phosphate
(McComb, Bowers and Posen, 1979).
The Michaelis–Menten equation for enzyme kinetics (Murray, 2002) predicts an initially high rate of
enzyme activity as the concentration of the substrate is relatively high, followed by a gradual
reduction in enzyme activity as the substrate is consumed. The enzyme alkaline phosphatase is also
known to be subject to competitive inhibition by inorganic phosphates via the formation of the
phospho–enzyme, thus blocking the active site to pPNP for dephosphorylation (Kim and Wyckoff,
1991).
The enzyme kinetics of alkaline phosphatase on this chemogenic substrate is the focus of this study,
in an attempt to determine the trend in the rate at which pNP concentration changes over time and
the impact of a high concentration of the inorganic phosphate product on enzyme activity. It is
hypothesised that the rate of enzyme activity will initially be

Bovine Blood Lab
For this experiment, whole bovine blood was used. The first process was to separate the blood into
cellular and plasma fractions. 100 µL of whole bovine blood was transferred into a yellow
microcentrifuge tube that was labeled WB using a P–200. 50 µL of whole blood was added to a blue
microcentrifuge tube labeled WB. Both tubes were capped and placed in ice. 2 mL of the remaining
blood was transferred into a Clear 2 mL tube using a P–2000 and centrifuged for 5 minutes at 8000
RPM. Afterward, 800 µL of the supernatant from the Clear tube was transferred into a yellow tube
labeled WP and 50 µL of the supernatant was added to a blue tube labeled WP. These tubes were
then capped and kept on ice.
Next, the plasma proteins from the blood were isolated ... Show more content on Helpwriting.net ...
A BSA standard serial dilution was created in wells A1–F1. Fifty microliters of dH20 were added to
each well and fifty microliters of the BSA stock was added to A1 and mixed. Fifty microliters from
A1 was transferred to B1 and this was repeated for C1, D1, and E1. In wells A2–A4, triplicates were
created by transferring 10 microliters from A1. Triplicates were created for the other BSA standards
using the same amount, but with a different tip for each standard. The samples WP, WB, Es, and Ap
(yellow tubes) were also used in the Bradford Assay. The dilutions for the samples were 1:10, 1:100,
1:500, and 1:1000. The first dilution consisted of 90 microliters of water and 10 microliters of the
sample. The second dilution consisted of 90 microliters of water and 10 microliters of the sample as
well. The third dilution composed of 80 microliters of water and 20 microliters of the sample. The
last dilution composed of 50 microliters of water and 50 microliters of the sample. These dilutions
were used for each sample and triplicates were generated for each dilution. Two hundred microliters
were added to each triplicated well for all the samples, including the standard. Then, the microplate
was placed on the spectrophotometer and the absorbance was

Bradford Assay On Unknown Concentrations Of Proteins
Bradford Assay on Unknown Concentrations of Proteins
Taylor Coleman
September 27, 2016
Lab Group 3
BIOL 1111: General Biology Lab
Fall 2015
Section 107
Chad Perry
Abstract Colorimetric assay is a process of determining a concentration of a solution based on
absorbance of light. The purpose of this lab is to determine if the Bradford assay is an accurate way
to determine an unknown concentration of two samples of protein. The Bradford assay is done by
measuring wavelength of light passing through a cuvette filled with Bradford dye and
concentrations of PBS and proteins. After the cuvettes are mixed they are placed into a
spectrophotometer to measure wavelength. The wavelength given will be used to plot a standard
curve based on concentration (x–axis) and wavelength (y–axis). The standard curve is then used to
measure an educated guess on the concentrations of unknown protein concentrations. We
hypothesized that if we use the Bradford assay and colorimetric spectrophotometry we can
determine an accurate concentration of two unknown concentrations of proteins. The results of this
lab failed to reject our hypothesis based on accurate measurements of protein concentrations. The
standard curves are drawn with a linear increasing slope. The Bradford assay is an accurate way to
demine the concentration of an unknown concentration.
Introduction Colorimetric assay is a process determining the concentration of a chemical element or
compound in a solution

Clinical Signs Of Bvdv Infection
Laboratory diagnosis The clinical signs of BVDV infection are highly variable and there are many
factors that affect severity of infection like viral strain, age of affected animals, immune status,
reproductive stage and mixed infection. Therefore, diagnosis based on history, clinical signs and
postmortem examination should be considered only as preliminary diagnosis. The accurate and
confirmatory diagnosis of BVDV infection depends only on laboratory diagnosis. 1– Virus isolation
BVDV isolation in cell culture, followed by identification, is considered as the most reliable method
for diagnosis. Many different clinical samples are suitable for BVDV isolation during the viremia
stage as nasal discharges, peripheral blood leukocytes, serum, semen, aborted fetuses and feces.
However, presence of maternal antibodies in serum and buffy coat samples from newly born
animals may neutralize the virus and decrease the sensitivity of virus isolation. Virus isolation
methods are labor intensive and may take several weeks. Furthermore, virus isolation cannot
differentiate between PI animals and transiently infected animals. The FBS used as supplement for
propagation of cell lines should be tested for BVDV contamination before usage to avoid false
positive results. Dubovi et al. 2013 stated that there are many factors that may affect the success in
BVDV isolation and discussed the misconception that BVDV isolation is the gold standard for
diagnosis. Some of those factors are variation

Disadvantages Of Fenugreek Seeds
Fenugreek seeds were bought from a local herbalist. The seeds were ground up and five different
solvents were used: chloroform, 70% ethanol, ethyl acetate, hexane and water. For each solvent, 2.5
grams of coarsely powdered seed material was extracted by refluxing with 25 ml of each solvent
stated above for a period of thirty minutes and kept overnight at room temperature before filtration.
The following day, after filtration, the solvents were evaporated until dryness using a rotary
evaporator and the crude extracts were weighed. To make a stock solution that could be used for the
subsequent experiments, five millimeters of dimethyl sulfoxide (DMSO) was added and two
hundred milligrams of the crude extract were weighed. Throughout the entire experiment, the
percentage of DMSO was kept constant (1%). MCF7 breast cancer cells were acquired from the
American Type Culture Collection (ATCC® HTB–22™). The media was supplemented with 10 %
(v/v) heat–inactivated fetal bovine serum (FBS) (Gibco Invitrogen), antibiotics 100 U/ml penicillin
(Lonza) and 100 µg/ml streptomycin (Lonza), 2 mM L–glutamine (Lonza) in addition to 25µM 4–
(2–hydroxyethyl)–1 piperazineethanesulfonic acid (HEPES) (Biowest, France). Cells were
maintained at 37 °C, 5% CO2 humidified incubator. The cell viability assay was performed using
the 3–(4,5–dimethyl–2–thiazoyl)–2,5–diphenyl–2H–tetrazolium bromide (MTT) as described
previously (Mosmann, 1983), with minor modification. In summary, 7×103 cells were seeded

Absorbance Values
Abstract:
Claims were made that manufacturers had improperly labeled protein concentrations on their
products. Concentrations of these three different protein–containing liquids, whole milk, soy milk,
and a protein solution were determined using a Bradford dye reagent and a spectrophotometer to
measure the absorbance of each. Absorbance of a set of standard known protein concentrations from
a range of 0.125 mg/ml to 2.000 mg/ml were measured using the spectrophotometer at a set
wavelength and this data was plotted. The equation generated from the best fit line of this data was
used to determine the unknown concentrations of the three different samples. These values were
then compared to the protein concentrations obtained from the corresponding ... Show more content
This would indicate that either the food labels or the calculated data are incorrect. The data was
dependent on a linear relationship between absorbance and protein concentration. However, the
linear relationship was based on only seven data points, two of which were omitted due to
inconsistency, and was only representative up to 2.000 mg/ml protein concentration. Because the
data sample was small and covered a small range of concentrations, the food label concentrations
cannot be determined as inaccurate. The more likely cause would be that the linearity assumption of
the correlation between the absorbance and the protein concentration is

Electrospinning Essay
Electrospinning is a convenient method for fabricating various nanofibrous scaffolds for biomedical
applications. In electrospinning process, a conductive collector device plays a critical role in
determining the shape and the structure of the nanofibrous scaffold; however, the preparation of the
collector is often complex. In this study, a novel, flexible, and inexpensive approach based on
pencil–on–paper method was developed for preparing collectors used in electrospinning. Graphite is
a conductive material widely used as a component in the pencil core. A series of collectors were
prepared through drawing different patterns on folded and unfolded paper using a pencil, followed
by successfully fabricating two–dimensional (2D) nanofibrous ... Show more content on
Helpwriting.net ...
We believe that the method developed in this study would provide a convenient approach to
fabricate complex collectors for producing ordered 2D or 3D nanofibrous structures.
2. Materials and methods
2.1. Electrospinning
The collector was fabricated using a common 5B pencil and A4 paper. Paper was tailored to suitable
size, and different patterns were drawn using a pencil. Subsequently, the paper was folded in
different shapes as collectors. Poly (ε–caprolactone) (PCL, MW = 60,000) was dissolved in
dichloromethane/ethanol (v:v =3:1) to form a homogeneous solution with a concentration of 15
wt%, and polyvinyl pyrrolidone (PVP, MW = 1,300,000) was dissolved in ethanol at a concentration
of 8 wt%. After that, the solution was transferred to a syringe fastened on an infusion pump. The
solution was pumped at a rate of 0.5 mL/h. 8 kV voltage was applied between the needle and the
collecter placed 10 cm away to form an electrospinning jet. The collectors and fibers were observed
using a camera (Nikon, Japan) and scanning electron microscopy (SEM) (JSM–7011F, Japan).
2.2. Cell experiment
Murine calvarial preosteoblasts (MC3T3–E1) were selected as the model cell to evaluate the
cytotoxicity of the electrospun fibers. The nanofibrous membranes and films after sterilization under
UV radiation were cut into small pieces, placed in a cell culture plate, and soaked in the culture
medium (DMEM/F–12

Lab Report : Bca Protein Assay
Shrutika Mintri/ E2 Date : 09/17/2014 Lab Report #1 Project Title: BCA Protein Assay Purpose:
BCA (Bicinchoninic acid) Assay otherwise known as the "Smith Assay" has the fundamental
purpose of determining the protein concentration of the two given unknown proteins in the sample
solutions. The absorbance is measured using a Plate reader and a Standard curve is generated. Also,
the different types of pipetting techniques are assessed in this Assay. Methods: (1) There are two
methods using which the BCA Protein Assay can be performed– Test tube method and Micro plate
method. The following experiment has been performed using the Microplate procedure. Vial Volume
of Diluent Volume and Source of BSA Final BSA Concentration A 325 μL 325μL of Stock
1000μg/ml B 325μL 325μL of vial A dilution 500μg/ml C 325μL 325μL of vial B dilution 250μg/ml
D 325μL 325μL of vial C dilution 125μg/ml E 400μL 0 0μg/ml= Blank 1. Preparation of Diluted
Albumin (BSA) Standards: The following Diluted Albumin (BSA) Standards have been prepared
using Bovine Serum Albumin (BSA) Standard Solution (2.0mg/ml) and the Diluent Phosphate
Buffered Saline (PBS) having

Protein Concentration Lab Report
Figure 3. Bradford Standard Curve using 10mg/ml BSA Protein. 10mg/ml BSA protein stock was
serially diluted to cover a 5–2000µg/mL concentration range. 50 µL of dilutions were added to
1.5mL Bradford Reagent, left at room temperature for 5minutes, and assayed at 595nm, against a
blank of 50 µL DI H20 and 1.5mL Bradford Reagent.1
________________________________________ From the Bradford standard curve equation, the
protein concentration of the crude lysate was calculated to be 0.1402 µg/mL, and the protein
concentration in Fractions 1, 2 and 3 was found to be 11.5038 µg/mL, 11.5038 µg/mL, and 26.6553
µg/mL, respectively. Although Fraction 2 appeared to have the most intense yellow coloring,
Fraction 3 had the highest protein concentration. ... Show more content on Helpwriting.net ...
The Stokes radius of a molecule is the effective radius a molecule has as it moves through a
solution. Molecular weight of a molecule is sometimes correlated with the Stokes radius of a
molecule, and thus the elution trend in gel–filtration, but this is not without exceptions. Thus, Stokes
radius is a better predictor, because although two molecules may have the same molecular weight, a
longer, more branched molecule will have a larger Stokes radius than a smaller molecule.1 In the
separation of blue dextran, cytochrome c, and potassium chromate, the molecular weight of the
molecules were proportionate to their molecular sizes. The largest molecule, blue dextran, eluted
first, followed by cytochrome c, and then the smallest molecule potassium chromate. Blue dextran
has the largest molecular weight, and potassium chromate has the smallest. The results for elution
order are in accordance with what the theoretical, predicted results were. The efficiency of the
separation was not as ideal as would be predicted on the basis of molecular weights. The molecular
weights of the molecules are 3 orders of magnitude different, and should produce well–resolved
peaks.1 Experimental visual results show peak overlapping around 2mL and 3mL, as the colors
were mixed at these elution volumes. This peak overlapping may be attributed to peak broadening.
Peak broadening has many causes, but of particular

Advantages And Disadvantages Of Cdna
troubleshooting. As the control RNA also did not yield any results through the several runs
attempted, it is still unknown why long template cDNA was not reliably produced for the next step
of the sequencing process. It is possible further optimisation is required for cDNA to be synthesised,
but funding and time constraints limited it for this study.
Further studies in this area may use the same reagents and protocols used with further optimisation
to yield results. Other possibilities are attempting different primers to try and create cDNA that
covers the whole genome.
4.7 Conclusions
The aims of the study were to develop molecular assays to detect and type EV from patient samples
and describe the virus distribution and epidemiology in South Africa. A single EV real–time ... Show
more content on Helpwriting.net ...
The advantage of the simplex, is that it is more robust and more sensitive that multiplex PCRs due
to the nature of few primers and probes in the assay. The disadvantage is that the sequencing assay
to be based on these results needs to then amplify all EV, instead of more specifically a species per
assay. Nix et al developed an assay for this purpose in 2006 (Nix et al., 2006). This assay was able
to detect all EV serotypes spread across all 4 HEV species. In this study, more recently classified EV
were detected by this assay: EV99, EV102, and EV114. These are little described viruses with no
clear disease association. The epidemiology of EV in South Africa showed a general concordance
with other studies (Benschop et al., 2010, Apostol et al., 2012, Dhole et al., 2009). HEV–B and
HEV–C contained the most number of typed viruses in the country, though as mentioned, many
previous studies were biased due to the use of culture before virus typing, instead of typing directly
from patient samples. Echovirus 30 is seen to be the predominant serotype in Europe, but elsewhere
in the world, the viral distribution is more even, with serotypes co–circulating (Oberste

Identifying An Organism With A Biological Lens Essay
Spectrophotometrically Identifying Alfalfa Sprouts' Chloroplasts and Lysosomes Isolated in a
Sucrose Gradient
Introduction: Conceptualizing an organism with a biological lens involves understanding the
organism's components and how their individual functions characterize the overarching entity. For a
body, this means understanding organs and tissues. For a cell, this means classifying its organelles
and discerning their functions. Analysis of subcellular components has become a major focus in
biochemical studies, and it relies heavily on the fractionation of organelles. Fractioning and isolating
organelles essentially enables a scientist to elucidate organelle functions, which, as mentioned
before, adds definition to the all–encompassing image of a cell. Modern studies on this topic tend to
target organelles with somewhat ambiguous functions, like anammoxosomes and melanosomes. By
isolating these subcellular components and analyzing them, biologists have unearthed
ammoxosomes' role in filling a large part of the Earth's atmosphere with nitrogen gas (Neumann et.
al, 2014). In addition to this, other biological organizations have managed to sequester intact
melanosomes from cells, separating them from other items of similar densities, and study the
activity of V–Type ATPase proteins while they are in a subcellular membrane (Pelkonen et. al,
2016). The general procedure for executing an experiment like these involves lysing cells,
centrifuging their components, and separating

Benefits Of Prunus Mume Fruit
Basic functions of prunus mume fruit inside the body
 The Citric acid present in the plum fruit helps in the absorption of vitamins and enzymes, thus
preventing several diseases and reduces fatigue.
 Eating plum fruit on a regular basis can stimulate the secretion of parotid gland hormone, which
can prevent aging process in humans.
 Prunus mume intake increases metabolism and aids in the process of body's energy restoration
functions in a natural manner
How the prunus mume fruit should be used
 Although the Chinese Plum is edible, it should be taken with proper care for the desired effects.
Below are some of the points to keep in mind while consuming this fruit–
 The plum fruit should not be consumed by anyone suffering from cold or ... Show more content
Additionally, they help break down the harmful buildup of toxic fats in the body system and aids in
the process of purification of the body.
Benefits of Brown Rice Vinegar for the body
Among the several benefits of Brown Rice Vinegar for human body, the major ones are–
Slower the process of Ageing: The authentic brown rice vinegar contains approximately 20 amino
acids and 16 organic acids that help prevent the formation of toxic fat peroxides which generally
contributes to the process of aging.
Maintains the cholesterol level for a healthy heart: Since it inhibits fatty peroxides, brown rice
vinegar is known to help in the process of reducing or preventing the buildup of harmful cholesterol
on blood vessel walls thus helping in lowering the blood pressure, reducing the LDL cholesterol
levels and keeps the heart in better condition.
Improves the digestive health: The acetic acid contents of the brown rice vinegar helps in the
absorption of essential vitamins and minerals in the body thus improving the overall digestive
health.
Basic functions of Brown Rice Vinegar inside the

Material And Methods : Bm Mscs Cultivation
Material and Methods: BM–MSCs cultivation: BM–MSCs were obtained from Institute Pasture of
Iran and were cultured in Low Glucose Dulbecco 's modified Eagle 's medium (Low Glc–DMEM)
(Gibco Invitrogen, Karlsruhe, Germany) with 10% FBS (Gibco Invitrogen, Karlsruhe, Germany)
and 1% penicillin and streptomycin (Sigma–Aldrich, Schnelldorf, Germany). Then, BM–MSCs
were incubated in humified incubator at 37˚C and 5% CO2. Cell's medium was replaced every two
days. U266 cell line: Confirmed U266 cells, a human myeloma cell line, were purchased from
Institute Pasture of Iran. U266 cells were grown in RPMI–1640 (Gibco Invitrogen, Karlsruhe,
Germany) with 10% FBS (Gibco Invitrogen, Karlsruhe, Germany) and 1% penicillin and
streptomycin (Sigma–Aldrich, Schnelldorf, Germany) in T25 flask culture (Orange Scientific,
Braine–l 'Alleud, Belgium). Then, U266 cells incubated in humified incubator at 37˚C and 5% CO2.
The medium was changed every day during all steps of experiments and Trypan blue staining was
used for viability analyzing. BM–MSCs derived C.M preparation: The BM–MSCs were cultured in
T25 flask culture (Orange Scientific, Braine–l 'Alleud, Belgium) with DMEM medium and
incubated for reaching to 60–70% confluency. In the next step, DMEM medium was replaced with
RPMI–1640 medium without FBS and was incubated for 72 hours (hrs). After incubation; medium
was collected, cells and debris were removed by centrifugation. Supernatant was concentrated by 10
kDa MW cut–off ultrafiltration

Production of Crude Extract
Crude Extract
In order to produce crude extract, bovine tissue was obtained, precisely minced to exclude extra fat,
and then blended with pH 7.2 phosphate buffer. The purpose of blending the tissue with the buffer
was to pulverize the cells, causing them to release their contents evenly–most importantly lactate
dehydrogenase (LDH)–into the solution. Since other cell components such as proteases which
reduce LDH were also released from the lysed cells, the slurry was kept on ice to minimize their
kinetic activity. The solution was then centrifuged, creating a pellet made of cellular debris and
leaving behind the crude product in the supernatant. Using data from the enzyme assay, the crude
extract revealed 155±7 mg total protein, 4980±80 units of total activity, 32±2 units/mg of specific
activity, a 100% yield, and a purification factor of 1 fold (Table 1).
40% Supernatant
The purpose of the 40% ammonium sulfate fractionation step is to extract impurities with low
solubility–such as lipids–from the solution. Because ammonium sulfate salt is extremely soluble, it
hydrates competitively against weakly soluble impurities causing them to precipitate out of the
solution as a pellet. Hence, LDH is left behind in the 40% supernatant. This step revealed a slight
drop in total protein amount from 155±7 to 124±5 mg and comparatively no change in total activity
from 4980±80 to 4900±100 units (Table 1). This data agrees with the salting out theory which states
that since only weakly

Curcuma Longa Essay
RESULTS AND DISCUSSION
Qualitative Analysis
Results of Phytochemical Evaluation for Curcuma Longa Extract
The results of phytochemical evaluation of Curcuma Longa extract summarized in table 2.
Table 2: Shows the results of phytochemical evaluation of Curcuma Longa extract.
1 Alkaloids +
2 Tannins +
3 Glycosides +
4 Saponins +
5 Flavonoids +
6 Terpenoids +
7 Phenol +
Figure 3: Illustrates the results of phytochemical tests of Curcuma Longa extract.
The previous table 2 and figure 3 show the results of applied phytochemical tests for constituents of
Curcuma Longa extract which were all positive. The results indicate the presence of the active
constituents of curcuminoids. It also indicates the presence of alkaloids, tannins, phenolic
compounds, terpenoids, saponins and flavonoids. These phytochemicals were also reported in ...
Show more content on Helpwriting.net ...
No. Concentration μg/ml Absorbance
1 10 0.18
2 20 0.23
3 30 0.278
4 40 0.33
5 50 0.38
Concentration of Curcuma Longa extract in mcg/ml
Figure 5: Illustrates calibration curve of standard curcumin.
From the result in table 6 and figure 5 which illustrated the calibration curve of the UV
spectrophotometer with working standard Curcuma Longa extract and the r2 = 0.9999.
Curcumin in Gel Formulation

Table 7: Shows linearity of curcumin in the gel formulation.
No. Concentration μg/ml Absorbance
1 10 0.127
2 20 0.148
3 30 0.166
4 40 0.185
5 50 0.206
Concentration of Curcuma Longa extract in mcg/ml
Figure 6: Illustrates the curve of curcumin in the gel formulation. Assay Test of Curcumin Content
in Gel Formula
Assay of curcumin in gel formula was calculated using the following formula
Assay of curcumin = (absorbance of sample / absorbance of standard × concentration of standard /
concentration of sample) 100
= (0.127/0.180 × 10/10) 100 =

The Effects Of A Putative Anti Cancer Drug On Hela Cells
Introduction
Background
According to the Office of National Statistics, in 2013, 29% of deaths in the UK were cancer
related, making it the leading broad cause of death in the population. In the United States, 1 in 4
deaths are cancer related, and it is estimated that in a lifetime, there is 44% chance of contracting an
invasive cancer if you are male, and a 38% chance if you are female (Siegel et al. 2014). So,
although our knowledge and education into different cancer mutagens such as UV radiation,
chemical carcinogens and viruses (Alberts et al. 2002), as well as cancer physiology and treatments
may have improved, leading to a decrease in cancer related mortalities, there are still huge advances
to be had in the field (CRUK 2013). ... Show more content on Helpwriting.net ...
In another review paper in Nature, Masters (2000) described how cancer cell lines are usually from
fast growing, undifferentiated cancers that are easily transported and proliferated. HeLa cells may be
differentiated cervical carcinoma cells but they attain their ability to proliferate uncontrollably from
the protein HPV–18, which, combined with their ability to divide indefinitely, due to their
chromosomes not shortening on replication, mean that they are the perfect cell line to grow in vitro
(Watson 2010). Therefore, the HeLa model was used worldwide in research for not only cancers and
development of different chemotherapeutic drugs such as Cannabinoids (Guzman 2003), but also in
the research of genomics and vaccines including that for polio. It should also be noted, that the
conditions in a laboratory lend themselves to a very minimal level of genetic drift and combined
with control measures that assure that new cells are used every few weeks, the HeLa cell line is
actually very similar to that of the original group of cells (Masters 2000). These characteristics mean
that HeLa cells are an ideal cell line to use in this experiment.
In this experiment, analysis of cell viability will be achieved through an MTT assay. The salt 3–
(4,5–dimethyl–2–thiazolyl)–2,5–diphenyltetrazolium bromide (MTT) is often used in cancer drug
screening (Vistica et al. 1991) as it is

The Consequences Of Alcohol Analysis
Alcohol is a very widely used drug, and is therefore present in many clinical and forensic toxicology
cases. Quantitation of alcohol levels in patients affects the clinical treatment they receive as well as
any legal consequences associated with the incident that led to their hospitalization. Alcohol analysis
and its consequences for postmortem cases also forms a large part of the forensic toxicologist's
workload [1–2].
Although gas chromatography–mass spectrometry (GCMS) or gas chromatography flame ionization
detection (GC–FID) is the most accurate and reliable method of determining blood alcohol
concentration (BAC), it is expensive, time–consuming, and requires technical proficiency manhours
[1–2]. Therefore, BAC determination in a ... Show more content on Helpwriting.net ...
However, there is still contention over its existence, prevalence, and severity; this is evidenced by
recent articles and court cases contesting its relevance [3,7] . It is problematic that almost every
mention of this phenomenon goes back to a single study in 1995 that failed to put the issue to rest
[6].
Forensically, this research has multiple implications. The presence and level of alcohol in a victim or
perpetrator's system has great bearing on their behavior and frame of mind. As a result, the amount
of alcohol in a person's blood can serve as a strong piece of evidence. From a sentencing standpoint,
the level of legal consequences assigned to an individual is also be dependent on the amount of
intoxication detected [1]. With these consequences in mind, it is imperative that the test used to
measure ethanol levels is reliable and accurate. If there is a possibility that the enzymatic assays
used widely throughout hospitals can be inaccurate under certain conditions, it is necessary that the
users and interpreters of those assays know that interference can occur and when it might happen. If
markers can be developed to determine when further testing for ethanol should be done, the
reliability of the test will be greatly increased. In this study, the aim will be to determine if lactate
and LDH in clinically possible antemortem concentrations can cause a significant

Lab Report
4.2 Materials and Method
4.2.1 Materials
The size of the particle should be large enough to allow three proteins coupling together onto one
particle. Therefore, Dynal® MyOneTM 1–μm carboxyl particle (Invitrogen®) and Merck® EM1–
100/40 particle were used as the carrier particles. If positive results were seen, the work would be
expanded to particles in other sizes, materials and manufacturers, such as Allrun® PM3–050 or
Dynal® M270 particles.
The proteins chosen for coupling were mouse IgG, streptavidin and APC. They carry different
functions and could give a good reflection of the possible functional proteins used in future particle
application. Mouse IgG (Lampire®) stock solution was filtered with a 0.45μm filter before diluting
into ... Show more content on Helpwriting.net ...
For quality control purpose, the CS&T particles were also running daily before testing the samples
to re–set the configuration. The machine was serviced and maintained routinely and worked in good
condition.
96–well U–bottom white polypropylene plate (Grenier®) was used for fluorophore loading assay
and particle washing. The 1.5ml polypropylene reaction tubes (Grenier®) were used for general
particle treatment and protein coupling. Also, tube magnet and plate magnet were used in particle
separation. All the consumables were routinely used in Anteo and they were proven suitable for
treating magnetic particles.
4.2.2 Method
The experiment in this chapter was divided into two parts. First, the particles were coupled with
signal proteins individually at different coupling concentration, and a titration curve was generated
to give supportive information for later co–coupling experiments, including the gate setting and
spectrum compensation values. Second, the particles were co–coupled with three proteins at
different protein coupling concentration level. Under different concentration combination, each
protein would show different signal response and it help to determine the optimal loading
concentration used in multifunctional particle construction.
The main particle used in this experiment was Dynal® MyOneTM

Bradford And Lowry Assay Lab Report
The Bradford and Lowry assay are two color changing proteins with similar processes. Color
changes of the solution can be measured using a device called a spectrometer. In lab, the two assays
can be used to determine the amount of protein in a solution. This can be done in an experiment by
reacting a specific reagent with amino acids or peptide bonds present in the proteins. This produces
a color change in the solution depending on the amount of peptide or amino acid reactions with the
reagents. This color change can then be measured using a device called a spectrophotometer.
The Bradford assay is a one– step process, which involves the Coomassie dye reacting with the
amino acids arginine and lysine. It under goes changings in acidic conditions ... Show more content
The type of amino acid sample affected the ability of the reagents to react with the peptide bonds
and amino acids. It also affected the amount of amino acids and peptide bonds in the protein. Each
of the four proteins used in the experiment had the same concentration, which was reflected in the
accuracy of the standards. In relation to the Bradford reagent, the amount of reactions that occurred
depended on the amount of arginine and lysine amino acids present in the protein. In relation to the
Lowry reagent, the amount of reactions that occurred depended on the amount of tryptophan and
tyrosine amino acids in the protein and the number of peptide bonds linking the amino acids. The
absorbance measured in the spectrophotometer was affected also by the differences in amino acid
compositions and the ability of the reagents to react as a result. Since all the protein standards had
the same concentration of 0.5 mg/mL, it allowed for the accurate comparison of the absorbance,
mean, and standard deviation of each of the

Essay about Molecular Biology and Ldh
*Running title: Isolation and Characterization of Avian Lactate Dehydrogenase
To whom correspondence should be addressed Sylvia DaoudKinze and James Proestes, Department
of Biochemistry, Portland State University Professor, Portland Oregon, 97207–0751;
E–mail: sylviakinzie@gmail.com and proestoj@gmail.com
–––––––––––––––––––––––––––––––––––––––––––––––––
Keywords: Lactate, Dehydrogenase, Avian, Bradford Assay, Affinity Column
Background: Lactate Dehydrogenase also known as LDH is an important NADH dependent enzyme
in metabolism that catalyzes the conversion of Pyruvate to Lactate.
Results: Catalytic activity was detected from chicken breast muscle
Conclusion: Copious amount of catalytic activity was detected, indicating that ... Show more content
Then the total of 51.32 grams of chicken was weighed out, cut into small pieces and placed in a
blender along with 75 mL of cold extraction buffer.
Four short bursts (around 5 seconds per burst) were applied to complete the homogenization process
with a ten second pause between each pulse. The homogenized tissue solution was then poured into
three 50 mL centrifuge tubes, and was centrifuged at 15000 RPM for 20 minutes. The supernatant
poured through two layers of cheese cloth into a 50 mL falcon tube, and the volume was recorded.
Three 0.5 mL aliquots were saved.
Ammonium Sulfate Precipitation After centrifugation, the volume of each CFE was measured, and
then per every mL, .39 grams of ammonium sulfate was weighed out. The CFE was then placed in a
beaker that was suspended on top of ice in a larger beaker, which was placed on top of a stir plate.
Over a course of 20 minutes, ammonium sulfate was added to the CFE while stirring slowly. After
the completing adding all of the ammonium sulfate, solution was stirred for an addition 15 minutes.
Followed by centrifugation at 15000 RPM for 20 minutes.
Purification of LDH
Dialysis of the Protien Dialysis of the salted protein was completed by a lab T.A.
Affinity Column Chromatography To separate LDH from the rest of the protein, affinity column
chromatography method was used. A column with a plastic stopcock was loaded with Cibacron blue
affinity matrix by adding

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