Kenyatta university. dna extration docxLando Elvis
1. The document describes a HotSHOT DNA extraction method to isolate DNA from fish fins. The method uses sodium hydroxide (NaOH) to lyse cells and release DNA. The NaOH treatment denatures the DNA. Tris buffer is then used to neutralize the solution and renature the DNA.
2. The method involves homogenizing fish fins, treating with hot NaOH to lyse cells and denature DNA, cooling and neutralizing with Tris buffer. Centrifugation pellets debris and collects DNA in the supernatant.
3. The HotSHOT method provides a rapid, inexpensive way to obtain PCR-quality DNA from fish fins compared to traditional methods. The isolated DNA is suitable for
This document describes experiments to extract and analyze lipids from serum. In the first part, lipids were extracted from serum using chloroform-methanol and separated via centrifugation. The organic phase containing lipids was evaporated, redissolved in chloroform, and observed as a yellow liquid, indicating the presence of lipids. In the second part, thin layer chromatography was used to separate lipids in the extract on silica plates developed in two solvent systems, revealing at least two lipid components with different polarities. The experiments demonstrated techniques for lipid extraction and separation.
1. The document describes a laboratory experiment to estimate blood glucose levels using the glucose oxidase/Trinder's method. Glucose in blood samples is oxidized by glucose oxidase to produce hydrogen peroxide, which is measured colorimetrically.
2. Three blood samples were tested and their glucose concentrations calculated. Sample 3 had the highest concentration at 11.34 mmol/L, indicating hyperglycemia. Sample 2 was within the normal range, while Sample 1 was hypoglycemic at 2.75 mmol/L.
3. The results confirm the visual observation that Sample 3 appeared most colored, correctly identifying hyperglycemia in that sample based on the quantitative analysis. The method allows accurate glucose measurement to
This document describes the procedure for extracting RNA from yeast cells using the strong salt method and then identifying the three main components of RNA - phosphates, bases, and ribose. The key steps are:
1) Boiling yeast in a strong salt solution to lyse the cells and precipitate proteins, then centrifuging to separate RNA from cell debris.
2) Adjusting the pH of the supernatant to precipitate the RNA, then centrifuging and collecting the RNA precipitation.
3) Hydrolyzing the RNA with acid to break it into its three components, then using different reagents to identify each component through observed color changes.
The document summarizes an internship report on water quality analysis in different areas of Faisalabad, Pakistan. It discusses the importance of water quality analysis, parameters analyzed, methods used, and chain of custody procedures to ensure sample integrity. Key parameters analyzed include physical properties like taste, odor, color and turbidity, chemical properties like pH, hardness, metals and nutrients, and microbiological analysis of coliform bacteria. Standard methods are selected based on required precision, cost and water use. Proper sampling, preservation, labeling, sealing and documentation in a chain of custody form tracks samples from collection to analysis.
This document provides a summary of various laboratory methods for assessing water quality, including tests for inorganic constituents, hardness, arsenic, alkalinity, pH, biological contaminants, and filtration processes. The methods described include tests for fluoride, iron, sulfate, nitrate, and total dissolved solids. Procedures are outlined for assessing arsenic, alkalinity through titration, and testing for total coliform and E. coli bacteria using UV light and growth indicators. The final section briefly describes a water treatment filter house. The document serves as a training aid for learning water quality analysis techniques.
This document provides instructions for using TRIZOL Reagent to isolate total RNA from cells and tissues. TRIZOL Reagent uses phenol and guanidine isothiocyanate to simultaneously disrupt cells and dissolve cell components while maintaining RNA integrity. The reagent separates into aqueous and organic phases upon the addition of chloroform, allowing for RNA precipitation from the aqueous phase using isopropyl alcohol. Isolated RNA is free of protein and DNA contamination and can be used in various downstream applications. Proper precautions must be taken to prevent RNase contamination during the RNA isolation procedure.
Kenyatta university. dna extration docxLando Elvis
1. The document describes a HotSHOT DNA extraction method to isolate DNA from fish fins. The method uses sodium hydroxide (NaOH) to lyse cells and release DNA. The NaOH treatment denatures the DNA. Tris buffer is then used to neutralize the solution and renature the DNA.
2. The method involves homogenizing fish fins, treating with hot NaOH to lyse cells and denature DNA, cooling and neutralizing with Tris buffer. Centrifugation pellets debris and collects DNA in the supernatant.
3. The HotSHOT method provides a rapid, inexpensive way to obtain PCR-quality DNA from fish fins compared to traditional methods. The isolated DNA is suitable for
This document describes experiments to extract and analyze lipids from serum. In the first part, lipids were extracted from serum using chloroform-methanol and separated via centrifugation. The organic phase containing lipids was evaporated, redissolved in chloroform, and observed as a yellow liquid, indicating the presence of lipids. In the second part, thin layer chromatography was used to separate lipids in the extract on silica plates developed in two solvent systems, revealing at least two lipid components with different polarities. The experiments demonstrated techniques for lipid extraction and separation.
1. The document describes a laboratory experiment to estimate blood glucose levels using the glucose oxidase/Trinder's method. Glucose in blood samples is oxidized by glucose oxidase to produce hydrogen peroxide, which is measured colorimetrically.
2. Three blood samples were tested and their glucose concentrations calculated. Sample 3 had the highest concentration at 11.34 mmol/L, indicating hyperglycemia. Sample 2 was within the normal range, while Sample 1 was hypoglycemic at 2.75 mmol/L.
3. The results confirm the visual observation that Sample 3 appeared most colored, correctly identifying hyperglycemia in that sample based on the quantitative analysis. The method allows accurate glucose measurement to
This document describes the procedure for extracting RNA from yeast cells using the strong salt method and then identifying the three main components of RNA - phosphates, bases, and ribose. The key steps are:
1) Boiling yeast in a strong salt solution to lyse the cells and precipitate proteins, then centrifuging to separate RNA from cell debris.
2) Adjusting the pH of the supernatant to precipitate the RNA, then centrifuging and collecting the RNA precipitation.
3) Hydrolyzing the RNA with acid to break it into its three components, then using different reagents to identify each component through observed color changes.
The document summarizes an internship report on water quality analysis in different areas of Faisalabad, Pakistan. It discusses the importance of water quality analysis, parameters analyzed, methods used, and chain of custody procedures to ensure sample integrity. Key parameters analyzed include physical properties like taste, odor, color and turbidity, chemical properties like pH, hardness, metals and nutrients, and microbiological analysis of coliform bacteria. Standard methods are selected based on required precision, cost and water use. Proper sampling, preservation, labeling, sealing and documentation in a chain of custody form tracks samples from collection to analysis.
This document provides a summary of various laboratory methods for assessing water quality, including tests for inorganic constituents, hardness, arsenic, alkalinity, pH, biological contaminants, and filtration processes. The methods described include tests for fluoride, iron, sulfate, nitrate, and total dissolved solids. Procedures are outlined for assessing arsenic, alkalinity through titration, and testing for total coliform and E. coli bacteria using UV light and growth indicators. The final section briefly describes a water treatment filter house. The document serves as a training aid for learning water quality analysis techniques.
This document provides instructions for using TRIZOL Reagent to isolate total RNA from cells and tissues. TRIZOL Reagent uses phenol and guanidine isothiocyanate to simultaneously disrupt cells and dissolve cell components while maintaining RNA integrity. The reagent separates into aqueous and organic phases upon the addition of chloroform, allowing for RNA precipitation from the aqueous phase using isopropyl alcohol. Isolated RNA is free of protein and DNA contamination and can be used in various downstream applications. Proper precautions must be taken to prevent RNase contamination during the RNA isolation procedure.
This document summarizes a study on the pharmacokinetics and tissue distribution of 8-acetamino-isocorydine (AICD) in rats using HPLC-DAD analysis. The study developed and validated an HPLC-DAD method to analyze AICD levels in plasma and tissue samples. Rats were administered AICD intravenously or orally, and plasma and tissue samples were collected over time. The results showed AICD had half-lives of 2.2 hours and 2.0 hours in rats following intravenous and oral administration, respectively. Following intravenous dosing, high AICD concentrations were found in the lungs, kidneys and liver, suggesting it may be effective for
BioStar Lifetech Genomic DNA extraction kit(rbp96 b) protocolBioStar Lifetech
This document provides protocols for using a 96-well genomic DNA extraction kit to purify DNA from blood, tissues, and other samples in a high-throughput format. The kit uses proteinase K and chaotropic salts to lyse cells and degrade proteins, then binds DNA to a glass fiber plate. DNA is washed and eluted in less than an hour for use in downstream applications such as PCR. The document describes vacuum and spin column protocols for blood, tissues, and other sample types.
Presentation on increased bioavailability of breviscapine via a pluronic p85 ...2503jyoti
Breviscapine is a hydrophobic drug used in cerebovascular diseases.It's bioavailability due to it's efflux by P-gp system.pluronics are non-ionic tri-bolic co-polymers made up of central part of PPO(hydrophobic) which is flanked by two PEO(hydrophilic) molecules
This study validated a noninvasive method to measure steroid hormones from the holding water of zebrafish. The study found:
1) Cortisol levels in holding water correlated positively with plasma cortisol levels in zebrafish, allowing noninvasive measurement of circulating cortisol.
2) While 11-ketotestosterone (KT) levels did not correlate between holding water and plasma, KT levels showed a significant sex difference detectable in holding water, allowing noninvasive sex determination.
3) An ACTH challenge test showed that induced increases in plasma cortisol were reliably detected in increased holding water cortisol levels, demonstrating the method can detect changes in circulating hormone levels.
Biochemistry is a basic science which deals with chemical nature and chemical behaviour of living matter and with the reactions and processes they undergo.
Biochemistry involves the study of:
Chemical constituents of living matter.
Chemical changes which occur in the organism during digestion, absorption and excretion.
Chemical changes which occur during growth and multiplication of the organism.
Transformation of one form of chemical constituent to the other.
Energy changes involved in such transformation.
Note:- The term “Biochemistry” was first introduced by German chemist Carl Neuberg in 1903 from Greek word “bios” means “life”.
It is mainly deals with the biochemical aspects that are involved in several conditions.
The results of qualitative and quantitative analysis of body fluids assist the clinicians in the diagnosis, treatment and prevention of the disease and drug monitoring, tissue and organ transplantation, forensic investigations and so on.
Various biological fluids subjected to chemical tests and assays include blood, plasma, serum, urine, cerebrospinal fluid (CSF), ascetic fluid, pleural fluid, faeces, calculi and tissues.
Note:- Modern day medical practice is highly dependent on the laboratory analysis of body fluids, especially the blood. The disease manifestations are reflected in the composition of blood and other tissues.
Hence, the demarcation of abnormal from normal constituents of the body is another aim of the study of clinical biochemistry.
RNA isolation is the purification of RNA from biological samples. It requires strict precautions to avoid degradation by RNases, enzymes that naturally degrade RNA. The TRIzol method uses phenol and chloroform to separate RNA, DNA and proteins during cell lysis and homogenization. Chloroform separates the mixture into aqueous and organic phases, with RNA remaining in the aqueous phase. RNA is then precipitated with isopropanol and washed with ethanol before being re-dissolved in water. RNA concentration and purity is determined spectroscopically. Filter-based and magnetic particle methods provide alternative RNA isolation techniques.
Method Development and Validation of Tetracycline Antibiotics and their Epime...Dr. Mukesh Raikwar
This document describes the development and validation of a method to detect tetracycline antibiotics and their epimers in marine products according to EU guidelines. The method uses HPLC and LC/MS/MS to separate and detect 7 tetracyclines (tetracycline, oxytetracycline, doxytetracycline, chlortetracycline, and their epimers) in shrimp samples. Validation showed good linearity, recovery, specificity, and other parameters within EU limits. The method provides a simple way to reliably detect tetracycline residues in marine products at the MRL level set by the EU.
Jeevani ( Role of clinical biochemistry in laboratory)Potu Jeevani
The cell is considered the structural and functional unit of life. It contains several organelles that perform important functions. The nucleus contains dense bodies like the nucleolus that is rich in RNA. Mitochondria are known as the powerhouses of the cell and have inner and outer membranes with cristae to increase surface area. Other organelles include the endoplasmic reticulum for protein biosynthesis, Golgi apparatus for membrane synthesis, lysosomes for digestion, and peroxisomes that contain enzymes to protect the cell. Clinical biochemistry utilizes biological fluids and cells to diagnose diseases through quantitative and qualitative tests. Precise methods and quality controls are important for accurate results.
Nucleic acid diagnostic and removal technics from biopharmaceuticalsElahehEntezarmahdi
This document discusses various nucleic acid-based techniques used in biopharmaceuticals, including plasmid cloning, DNA extraction, gel electrophoresis, polymerase chain reaction (PCR), reverse transcription PCR (RT-PCR), real-time PCR, DNA hybridization probes, and methods for detecting pathogens like mycoplasma, malaria, and Trypanosoma cruzi. It also covers techniques for removing nucleic acids like precipitation using polyethyleneimine and treatment with nucleases like DNase I and Benzonase endonuclease.
The researchers aimed to purify cellular retinol binding protein (CRBP) from bovine liver. Through a process involving homogenization, centrifugation, cation exchange chromatography, gel filtration, and concentration, they obtained a final product. However, characterization through SDS-PAGE and absorption spectroscopy identified the protein as catalase rather than CRBP. Despite initial absorption at 350nm for CRBP, the maximal absorption and thermal/pH profiles matched those of catalase. The purification resulted in the isolation of catalase rather than the intended CRBP.
Bacterial total protein and membrane protein extractionCreative BioMart
This document describes methods for extracting total protein and membrane proteins from bacterial samples. It provides details on reagents needed, equipment, and procedures for a simple extraction method using lysis buffer, ultrasonication and centrifugation. It also details a method using Trizol lysis solution and subsequent protein precipitation steps. Finally, it outlines a Triton X-114 detergent method for extracting hydrophobic membrane proteins, involving cell washing, detergent phase separation at different temperatures, and acetone precipitation before analysis.
The document discusses the purpose and functions of clinical pathology and laboratory medicine in assisting clinicians. It aims to confirm or reject diagnoses, provide guidance in patient management, establish prognoses, detect diseases through screening and monitoring follow-up therapy. It also discusses the scope of clinical chemistry, including biochemistry, instrumentation, computers, pharmacology and more. Point-of-care testing is described as testing done near patients using portable analyzers.
Biological screening of herbal drugs for anti cancer activityshafna hussain
This document summarizes several methods for screening potential anti-cancer compounds in vitro and in vivo. It describes assays to test compounds' effects on cell viability, growth, and metabolism in cell cultures, including trypan blue dye uptake, [3H]thymidine uptake, and MTT dye conversion assays. For in vivo models, it mentions using chemically-induced cancer in rats to test compounds' effects on tumor doubling time and growth. Common carcinogens mentioned are dimethylhydrazine and 1-methyl-1-nitrosourea used to induce colorectal and breast cancers in rat models respectively.
Pesticide residue detection methods by making use of the quantum related technologies are described, the motivation is to push the detection limit, to protect the environment we are to survive beyond what Stephen Hawking predicted!
This document discusses the bacterial endotoxin test using the gel-clot method. It provides background on endotoxins, which are toxic lipopolysaccharides found in the cell walls of gram-negative bacteria. Three techniques can test for endotoxins - gel-clot, turbidimetric, and chromogenic. The gel-clot method uses amoebocyte lysate extracted from horseshoe crabs to detect endotoxins. The test involves incubating lysate with samples and checking for clot formation, which indicates the presence of endotoxins. Endotoxin limits are specified for injectable pharmaceuticals to ensure safety. The document outlines the bacterial endotoxin test procedure using the gel-
Erythrocytes, or red blood cells, have potential as carriers for drug delivery. They are biocompatible, biodegradable, have long circulation times, and can be loaded with drugs using various chemical and physical methods. Resealed erythrocytes are prepared by breaking open erythrocytes, loading them with drugs from solution, and resealing the cells. Various methods exist for drug loading, including hypotonic lysis, electroporation, and endocytosis. Loaded erythrocytes have applications in targeting drugs to tissues like the liver and spleen, and treating conditions like cancer, parasites, and iron overload. Further research continues to improve drug loading efficiency and stability of resealed
ALCOHOL WITHDRAWAL SYNDROME. Dr. LANDO ELVIS.pptxLando Elvis
This document discusses alcohol withdrawal syndrome (AWS) and its complications. It begins with an introduction to AWS and outlines its typical presentation and pathophysiology. It then describes several potential complications of AWS including alcohol withdrawal seizures, alcoholic hallucinosis, delirium tremens, and Wernicke's encephalopathy. For each complication, it discusses signs and symptoms, risk factors, prevention and management strategies. Throughout it emphasizes the importance of inpatient management and monitoring for AWS given the risk of serious complications.
1. Septic shock is a life-threatening condition that occurs when sepsis leads to dangerously low blood pressure and organ dysfunction. It is caused by an uncontrolled immune response to infection that damages the cardiovascular system.
2. Early and aggressive treatment of septic shock is crucial and involves rapid administration of broad-spectrum antibiotics, controlling the infection source, and fluid resuscitation along with vasopressors to maintain adequate blood pressure over the first six hours.
3. The goals of fluid resuscitation in septic shock are to restore adequate tissue perfusion as indicated by targets such as a mean arterial pressure over 65 mmHg, urine output over 0.5 ml/kg/hr, and normalization of lactate
This document summarizes a study on the pharmacokinetics and tissue distribution of 8-acetamino-isocorydine (AICD) in rats using HPLC-DAD analysis. The study developed and validated an HPLC-DAD method to analyze AICD levels in plasma and tissue samples. Rats were administered AICD intravenously or orally, and plasma and tissue samples were collected over time. The results showed AICD had half-lives of 2.2 hours and 2.0 hours in rats following intravenous and oral administration, respectively. Following intravenous dosing, high AICD concentrations were found in the lungs, kidneys and liver, suggesting it may be effective for
BioStar Lifetech Genomic DNA extraction kit(rbp96 b) protocolBioStar Lifetech
This document provides protocols for using a 96-well genomic DNA extraction kit to purify DNA from blood, tissues, and other samples in a high-throughput format. The kit uses proteinase K and chaotropic salts to lyse cells and degrade proteins, then binds DNA to a glass fiber plate. DNA is washed and eluted in less than an hour for use in downstream applications such as PCR. The document describes vacuum and spin column protocols for blood, tissues, and other sample types.
Presentation on increased bioavailability of breviscapine via a pluronic p85 ...2503jyoti
Breviscapine is a hydrophobic drug used in cerebovascular diseases.It's bioavailability due to it's efflux by P-gp system.pluronics are non-ionic tri-bolic co-polymers made up of central part of PPO(hydrophobic) which is flanked by two PEO(hydrophilic) molecules
This study validated a noninvasive method to measure steroid hormones from the holding water of zebrafish. The study found:
1) Cortisol levels in holding water correlated positively with plasma cortisol levels in zebrafish, allowing noninvasive measurement of circulating cortisol.
2) While 11-ketotestosterone (KT) levels did not correlate between holding water and plasma, KT levels showed a significant sex difference detectable in holding water, allowing noninvasive sex determination.
3) An ACTH challenge test showed that induced increases in plasma cortisol were reliably detected in increased holding water cortisol levels, demonstrating the method can detect changes in circulating hormone levels.
Biochemistry is a basic science which deals with chemical nature and chemical behaviour of living matter and with the reactions and processes they undergo.
Biochemistry involves the study of:
Chemical constituents of living matter.
Chemical changes which occur in the organism during digestion, absorption and excretion.
Chemical changes which occur during growth and multiplication of the organism.
Transformation of one form of chemical constituent to the other.
Energy changes involved in such transformation.
Note:- The term “Biochemistry” was first introduced by German chemist Carl Neuberg in 1903 from Greek word “bios” means “life”.
It is mainly deals with the biochemical aspects that are involved in several conditions.
The results of qualitative and quantitative analysis of body fluids assist the clinicians in the diagnosis, treatment and prevention of the disease and drug monitoring, tissue and organ transplantation, forensic investigations and so on.
Various biological fluids subjected to chemical tests and assays include blood, plasma, serum, urine, cerebrospinal fluid (CSF), ascetic fluid, pleural fluid, faeces, calculi and tissues.
Note:- Modern day medical practice is highly dependent on the laboratory analysis of body fluids, especially the blood. The disease manifestations are reflected in the composition of blood and other tissues.
Hence, the demarcation of abnormal from normal constituents of the body is another aim of the study of clinical biochemistry.
RNA isolation is the purification of RNA from biological samples. It requires strict precautions to avoid degradation by RNases, enzymes that naturally degrade RNA. The TRIzol method uses phenol and chloroform to separate RNA, DNA and proteins during cell lysis and homogenization. Chloroform separates the mixture into aqueous and organic phases, with RNA remaining in the aqueous phase. RNA is then precipitated with isopropanol and washed with ethanol before being re-dissolved in water. RNA concentration and purity is determined spectroscopically. Filter-based and magnetic particle methods provide alternative RNA isolation techniques.
Method Development and Validation of Tetracycline Antibiotics and their Epime...Dr. Mukesh Raikwar
This document describes the development and validation of a method to detect tetracycline antibiotics and their epimers in marine products according to EU guidelines. The method uses HPLC and LC/MS/MS to separate and detect 7 tetracyclines (tetracycline, oxytetracycline, doxytetracycline, chlortetracycline, and their epimers) in shrimp samples. Validation showed good linearity, recovery, specificity, and other parameters within EU limits. The method provides a simple way to reliably detect tetracycline residues in marine products at the MRL level set by the EU.
Jeevani ( Role of clinical biochemistry in laboratory)Potu Jeevani
The cell is considered the structural and functional unit of life. It contains several organelles that perform important functions. The nucleus contains dense bodies like the nucleolus that is rich in RNA. Mitochondria are known as the powerhouses of the cell and have inner and outer membranes with cristae to increase surface area. Other organelles include the endoplasmic reticulum for protein biosynthesis, Golgi apparatus for membrane synthesis, lysosomes for digestion, and peroxisomes that contain enzymes to protect the cell. Clinical biochemistry utilizes biological fluids and cells to diagnose diseases through quantitative and qualitative tests. Precise methods and quality controls are important for accurate results.
Nucleic acid diagnostic and removal technics from biopharmaceuticalsElahehEntezarmahdi
This document discusses various nucleic acid-based techniques used in biopharmaceuticals, including plasmid cloning, DNA extraction, gel electrophoresis, polymerase chain reaction (PCR), reverse transcription PCR (RT-PCR), real-time PCR, DNA hybridization probes, and methods for detecting pathogens like mycoplasma, malaria, and Trypanosoma cruzi. It also covers techniques for removing nucleic acids like precipitation using polyethyleneimine and treatment with nucleases like DNase I and Benzonase endonuclease.
The researchers aimed to purify cellular retinol binding protein (CRBP) from bovine liver. Through a process involving homogenization, centrifugation, cation exchange chromatography, gel filtration, and concentration, they obtained a final product. However, characterization through SDS-PAGE and absorption spectroscopy identified the protein as catalase rather than CRBP. Despite initial absorption at 350nm for CRBP, the maximal absorption and thermal/pH profiles matched those of catalase. The purification resulted in the isolation of catalase rather than the intended CRBP.
Bacterial total protein and membrane protein extractionCreative BioMart
This document describes methods for extracting total protein and membrane proteins from bacterial samples. It provides details on reagents needed, equipment, and procedures for a simple extraction method using lysis buffer, ultrasonication and centrifugation. It also details a method using Trizol lysis solution and subsequent protein precipitation steps. Finally, it outlines a Triton X-114 detergent method for extracting hydrophobic membrane proteins, involving cell washing, detergent phase separation at different temperatures, and acetone precipitation before analysis.
The document discusses the purpose and functions of clinical pathology and laboratory medicine in assisting clinicians. It aims to confirm or reject diagnoses, provide guidance in patient management, establish prognoses, detect diseases through screening and monitoring follow-up therapy. It also discusses the scope of clinical chemistry, including biochemistry, instrumentation, computers, pharmacology and more. Point-of-care testing is described as testing done near patients using portable analyzers.
Biological screening of herbal drugs for anti cancer activityshafna hussain
This document summarizes several methods for screening potential anti-cancer compounds in vitro and in vivo. It describes assays to test compounds' effects on cell viability, growth, and metabolism in cell cultures, including trypan blue dye uptake, [3H]thymidine uptake, and MTT dye conversion assays. For in vivo models, it mentions using chemically-induced cancer in rats to test compounds' effects on tumor doubling time and growth. Common carcinogens mentioned are dimethylhydrazine and 1-methyl-1-nitrosourea used to induce colorectal and breast cancers in rat models respectively.
Pesticide residue detection methods by making use of the quantum related technologies are described, the motivation is to push the detection limit, to protect the environment we are to survive beyond what Stephen Hawking predicted!
This document discusses the bacterial endotoxin test using the gel-clot method. It provides background on endotoxins, which are toxic lipopolysaccharides found in the cell walls of gram-negative bacteria. Three techniques can test for endotoxins - gel-clot, turbidimetric, and chromogenic. The gel-clot method uses amoebocyte lysate extracted from horseshoe crabs to detect endotoxins. The test involves incubating lysate with samples and checking for clot formation, which indicates the presence of endotoxins. Endotoxin limits are specified for injectable pharmaceuticals to ensure safety. The document outlines the bacterial endotoxin test procedure using the gel-
Erythrocytes, or red blood cells, have potential as carriers for drug delivery. They are biocompatible, biodegradable, have long circulation times, and can be loaded with drugs using various chemical and physical methods. Resealed erythrocytes are prepared by breaking open erythrocytes, loading them with drugs from solution, and resealing the cells. Various methods exist for drug loading, including hypotonic lysis, electroporation, and endocytosis. Loaded erythrocytes have applications in targeting drugs to tissues like the liver and spleen, and treating conditions like cancer, parasites, and iron overload. Further research continues to improve drug loading efficiency and stability of resealed
ALCOHOL WITHDRAWAL SYNDROME. Dr. LANDO ELVIS.pptxLando Elvis
This document discusses alcohol withdrawal syndrome (AWS) and its complications. It begins with an introduction to AWS and outlines its typical presentation and pathophysiology. It then describes several potential complications of AWS including alcohol withdrawal seizures, alcoholic hallucinosis, delirium tremens, and Wernicke's encephalopathy. For each complication, it discusses signs and symptoms, risk factors, prevention and management strategies. Throughout it emphasizes the importance of inpatient management and monitoring for AWS given the risk of serious complications.
1. Septic shock is a life-threatening condition that occurs when sepsis leads to dangerously low blood pressure and organ dysfunction. It is caused by an uncontrolled immune response to infection that damages the cardiovascular system.
2. Early and aggressive treatment of septic shock is crucial and involves rapid administration of broad-spectrum antibiotics, controlling the infection source, and fluid resuscitation along with vasopressors to maintain adequate blood pressure over the first six hours.
3. The goals of fluid resuscitation in septic shock are to restore adequate tissue perfusion as indicated by targets such as a mean arterial pressure over 65 mmHg, urine output over 0.5 ml/kg/hr, and normalization of lactate
Pyelonephritis is an inflammation of the kidney that is usually caused by a bacterial infection ascending from the bladder. It is more common in women due to their shorter urethra. Risk factors include urinary tract abnormalities, pregnancy, diabetes, and frequent sexual intercourse. Common symptoms are fever and flank pain. Diagnosis involves urine testing and culture. Treatment is usually oral fluoroquinolone antibiotics for a week. Chronic or recurring pyelonephritis can lead to kidney damage and renal failure if not properly treated.
EFFECTS OF PROLONGED FASTING IN PATIENTS.pptxLando Elvis
Prolonged fasting places the body in a state of metabolic stress. It goes through four stages as it adapts to using its stores of glycogen, fat, and protein for energy. Initially, it breaks down glycogen and enters gluconeogenesis. After 2-3 days of fasting, lipolysis increases to provide free fatty acids for beta-oxidation. Around 5 days of fasting, protein catabolism increases to provide amino acids for gluconeogenesis while growth hormone increases to preserve muscle mass. After 3 weeks without food, ketone bodies become the primary fuel for the brain. Prolonged starvation can damage vital organs and lead to deficiencies, dehydration, electrolyte imbalances, and eventually death.
Physiologic changes in pregnancy assighnment.pptxLando Elvis
Physiologic changes of pregnancy outlines the normal anatomical and physiological changes that occur during pregnancy to accommodate the growing fetus. There are significant cardiovascular, respiratory, hematologic, endocrine, urinary, gastrointestinal, integumentary, musculoskeletal, and reproductive system changes. The document discusses in detail the hormonal changes involved and their effects on various body systems. It emphasizes that these changes are normal adaptations to support pregnancy but may also increase risks for certain medical conditions if not properly managed.
The document discusses amputations, including definitions, indications, types, techniques, and rehabilitation. Amputation is the surgical removal of a limb or part of a limb. It may be necessary for conditions like peripheral vascular disease, trauma, burns, or malignant tumors. The main types are provisional, definitive end-bearing, and definitive non-end-bearing amputations. Proper pre-surgical evaluation, anesthesia use, soft tissue and bone cutting techniques, and post-operative care including physiotherapy and prosthetics are essential to optimize outcomes. Complications can include flap breakdown, gas gangrene, skin issues, ulceration, nerve damage, and phantom limb sensation.
Kenyatta university. serum uric acid docxLando Elvis
- The document describes a laboratory experiment to determine uric acid levels in serum or plasma samples using an enzymatic colorimetric method.
- Uric acid is produced from the breakdown of purines and circulates in plasma, where it is mainly excreted by the kidneys. Elevated levels can indicate renal insufficiency or conditions like gout.
- The enzymatic method involves uricase converting uric acid to allantoin with a color change reaction proportional to uric acid concentration, allowing samples to be quantified against a standard.
Kenyatta university. liver funtion testLando Elvis
This document reports on a practical experiment to determine liver function tests by measuring the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in a sample. The procedures, principles, normal ranges, and significance of ALT, AST, and their ratio are described. The results of the experiment found the ALT and AST levels to be within normal ranges, indicating normal liver function. Liver function tests provide information about liver health but must be interpreted in the full clinical context.
Kenyatta university. hmb201 dna repairdocxLando Elvis
Gene mutations can be hereditary or acquired. DNA has multiple repair mechanisms to correct damage from environmental factors and replication errors. These include mismatch repair, nucleotide excision repair, photoreactivation repair, base excision repair, and double-strand break repair through nonhomologous end joining or homologous recombination. Errors in DNA repair can lead to mutations and genetic disorders if damage is not corrected. The cell cycle is also regulated to check for DNA damage and delay progression until repairs are complete.
1. The determination of serum blood urea nitrogen (BUN) is a widely used screening test to evaluate kidney function. BUN levels provide information about prerenal, renal, and postrenal causes of hyperuremia.
2. The test measures the concentration of urea in the blood, which is produced from protein catabolism in the liver. Higher than normal BUN levels can indicate kidney dysfunction, dehydration, or a high-protein diet, while lower levels may be seen in liver disease or malnutrition.
3. For this test, the student measured a BUN level of 10.5 mg/dL or 1.7483 mmol/L, which is within the normal reference
Kenyatta university. atp muscle contractionLando Elvis
1. The experiment investigates how skeletal muscle maintains ATP levels during contraction by measuring the contraction of thin muscle strips when exposed to ATP, boiled ATP, or glucose solutions.
2. Muscle strips exposed to ATP solution contracted by an average of 2.5%, while those exposed to boiled ATP did not contract, indicating ATP is necessary to cause contraction.
3. Strips exposed to glucose solution contracted slightly, but the experiment could not determine if glucose provided energy for contraction since the muscle was dead tissue lacking metabolic pathways. The results show ATP is required for contraction but do not prove how contraction is energetically sustained in living muscle.
Kenyatta university. alkalyne phosphataseLando Elvis
1. The document reports on a test to measure alkaline phosphatase (ALP) levels in a serum sample. ALP is an enzyme found in various tissues including the liver and bone. Elevated ALP can indicate liver disease or bone disorders with increased bone cell activity.
2. The test measures the hydrolysis of p-Nitrophenyl phosphate to p-Nitrophenol, which has strong absorbance at 405nm. By measuring the change in absorbance over time, the ALP activity in the sample can be calculated.
3. The results of the test on the given sample show a normal, low level of ALP. This suggests the individual does not have liver disease or a bone
1. Albumin is a protein made by the liver that helps maintain fluid balance. Low albumin levels can indicate liver or kidney problems or excessive loss through urine.
2. This report details a procedure to estimate albumin levels in serum through binding with bromocresol green dye and measurement of absorbance. The student found a level of 27.82 mg/dl, slightly below the normal range.
3. Albumin levels provide information about liver function, nutrition, and calcium levels. Many factors can affect albumin so all must be considered when evaluating abnormal values, which can be caused by decreased synthesis, increased breakdown, or losses.
Kenyatta university. albumin estimation 2nd yr docxLando Elvis
- This document describes a laboratory experiment estimating albumin levels in serum using the dye-binding method with bromcresol green dye.
- Key points: Albumin strongly binds certain dyes like bromcresol green, forming complexes with distinct absorption spectra. This allows albumin concentration measurement despite excess dye. The student estimates an albumin level of 3.915 g/dL in a serum sample, within the normal reference range of 3.5-5.0 g/dL. Interpretation of albumin levels provides information about liver function, nutrition status, and calcium levels.
Kenyatta university, physiology pract 1Lando Elvis
The document discusses a practical report on a hematocrit blood test. It provides background information on what a hematocrit test measures and what it is used for. The test determines the percentage of red blood cells in the blood and is used to identify or monitor various blood-related conditions like anemia. The document then describes how the test was conducted on 24 participants, presenting their results in tables. It analyzes the data, finding the average hematocrit percentages were higher in males than females. Most participants fell within the normal ranges, though some females and males showed reduced hematocrit levels, indicating anemia.
Kenyatta university thyroid hormone synthesis cat1 taskLando Elvis
The document summarizes thyroid hormone synthesis. It discusses how iodine is transported into thyroid cells and used to iodinate tyrosine residues on thyroglobulin in the follicular lumen, catalyzed by thyroperoxidase. The iodinated tyrosines are then coupled to form the thyroid hormones T3 and T4. Factors that control hormone synthesis like iodine availability and TSH are described. Disorders of thyroid hormone synthesis including iodine deficiency, Graves' disease, and Hashimoto's thyroiditis are also summarized.
Kenyatta university ninhydrin prac assighnmentLando Elvis
The document describes a practical experiment using the ninhydrin screening test to detect amino acids. Ninhydrin reacts with amino acids in a pH range of 4-8, producing a purple colored product. The student conducted the test on solutions of arginine, tryptophan, tyrosine, cysteine, glycine, and glutamic acid. All solutions turned purple upon mixing with ninhydrin and heating for two minutes, except the distilled water control. The color change occurs because ninhydrin breaks down amino acids into aldehydes, ammonia, and carbon dioxide, then condenses with the breakdown products to form an intensely colored pigment.
Kenyatta university hemoglobin estimation Lando Elvis
This document reports on a practical experiment to determine hemoglobin levels using the cyanmethemoglobin method. The student provides background on hemoglobin and its complexes, describes the cyanmethemoglobin method and reagents, documents the procedures, observations and calculations, and analyzes the results. The student's hemoglobin level was increased, indicating polycythemia. The cyanmethemoglobin method provides an accurate means to assess anemia and guide transfusion decisions.
Kenyatta university epinephrine dedermination Lando Elvis
Epinephrine (adrenaline) is an important neurotransmitter and hormone. This report details a lab experiment detecting epinephrine using iron chloride, which causes a color change. Epinephrine was detected in one sample, shown by a blue-green then dark red color, but not in the control sample. The procedure demonstrates the characteristic chemical properties of epinephrine and how it can be identified.
This document discusses elephantiasis, also known as lymphatic filariasis, which is a disease prevalent in Kenya's coastal region. It is caused by parasitic roundworms transmitted by mosquitoes. The document provides background on the disease and risk factors for its spread. It then outlines a social work plan to address the increasing cases of elephantiasis in the community through awareness campaigns, meetings with authorities, community involvement in prevention and treatment efforts, and addressing stigma and misconceptions around the disease. Case studies are also presented to illustrate the impact of elephantiasis on individuals and challenges in controlling its spread.
2. Objectives
1. To isolate DNA fromfishfins.
2. To obtainPCR-qualityDNA fromthe fishfins.
BACKGROUND
DNA,the buildingblockof life isthe geneticmaterial forall livingorganismsanditcontains information
that iscrucial forheredity.DNA isolationisnecessaryforgeneticanalysisincludingscientific,medical,or
forensicpurposes.Presenceof lipids,proteins,polysaccharidesandsome otherimpuritiesinthe DNA
preparationcaninterfere withDNA analysismethodsbyreducingthe qualityof DNA, whichleadstoits
shorterstorage life.DNA canbe isolatedfromanylivingordeadorganism.Commonlyusedsourcesfor
DNA isolationare whole blood,hair,sperm, bones,nails,tissues,saliva, epithelialcells,urine, bacteria,
animal tissues,andplants.
The traditional methodsforthe isolationof DNA are more time consumingandthe reagentsusedare
costly.A fewmodificationsinthisprotocol make itsimple toprepare PCR-qualitygenomicDNA than
traditional methods.Inthismodifiedmethod(HotSHOTmethod),samplesare incubatedbrieflyinhot
NaOH andthenneutralizedbyTrisbuffer.Thismethodfoundtobe rapid,reliable,andinexpensive for
the isolationof PCR-qualityDNA fromzebrafish tissues.These advantagesmake ituseful forhigh-
throughput applications.Althoughitisaveryrapidand simple method,the qualityof DNA isnot
adequate forall applications.ItisfoundthatDNA obtainedfromthismethodisshearedandits
concentrationisveryless.The DNA isextensivelyshearedanditsconcentrationistoolow.Itisalso
possible thatthe DNA wasnot efficientlydigestedbythe restrictionenzyme becauseof interfering
substancesinthe DNA preparationorbecause the DNA didnot reanneal efficientlyafteritwas
neutralized.SothismethodismainlyforPCRrelatedapplications.AlthoughHotSHOTDNA preparation
islikelytobe limitedtoPCRapplications,the savingsintime andreagentsovertraditional DNA
preparationmethodsis substantial,andthe qualityof the resultsisasgoodas or slightlybetterthan
DNA preparedbytraditional methods.
Principle
The isolationof DNA usuallybeginswithlysisof tissue orcellswhichisessential forthe destructionof
proteinstructuresandalsoallowsthe release of nucleicacidsfromthe nucleus.Sodiumhydroxideis
mostlyusedtoextractDNA outof a cell.Ithelpsto breakdownthe cell wall bylooseningthe hard
structure of a cell wall ormembrane.More importantly,NaOHdisrupts the hydrogenbondingbetween
the Nitrogenbasesand convertingthe double-strandedDNA (dsDNA) (includingthe genomicDNA
(gDNA) andthe plasmid) tosingle strandedDNA (ssDNA).Thisprocessiscalleddenaturationandisthe
central part of thisDNA extractionprocedure.Presenceof sodiumhydroxidemakesthe solutionvery
3. basicor alkaline.Tris-Cl maintainsthe pHof the solution.Basicallyitreactswiththe lipopolysaccharides
presentonthe outermembrane whichmake the membrane permeable
Requirements
Fish fillet
Micropipette and pipette tips
Scalpel
Micro centrifuge tubes
Mortar and pestle
Centrifuge
Water bath
4o
c freezer
Reagents
50mM NaOH solution
1M Tris-Cl
Procedure
Pluck out 20-100 mg of fish fin using a sterile scalpel and a forceps.
Homogenize the fins using sterile mortar and pestle.
Aliquot 100 μl of 50 mM NaOH in to a microfuge tube.
Transfer the homogenized fins of zebra fish into this micro centrifuge tube.
Incubate at 95°C for 20min in a water bath.
After the incubation, the tubes are allowed to cool at 4°C.
Add 1/10th volume of 1 MTris-HCl (pH 8.0) for neutralizing the basic solution.
Centrifuge the sample at 5000rpm for 10min to pellet the debris. The supernatant obtained is
collected in afresh micro centrifuge tube (1-5μl of this supernatant can be used immediately
per 25 μL of PCR reaction mixture).
Store the sample for at least 3 months at 4°C or longer at -20°C freezer.
Observationsand result
-The exactpH is maintainedby- Tris-Cl.
-Denaturation isthe central partof thismethod.
4. -95°C is the incubationtemperatureforDenaturation.
-Neutralization isthe role of Tris-Cl.
-Cell denaturationisdone by- NaOH,itlyse cell toexpel nucleicmaterial,anddisrupt
nitrogenousbases.
DISCUSSIONAND CONCLUSION
Deoxyribonucleicacid (DNA) extractionisthe processbywhichDNA isseparatedfromproteins,
membranes,andothercellularmaterial containedinthe cell fromwhichitisrecovered .
Preparation DNA forPCR iscostlyand laborious,primarily becauseof the difficultyin physically
separatingDNA fromothertissue components.Thislevel of purification isunnecessaryformanyPCR
applications.A simplealternativeinthismethod,lysisinanalkaline reagent andneutralization witha
suitable buffer,hasbeen usedtoprepare genomicDNA from buccal swabs,whole blood,semenand
forensicsamplescollectedfromhumans.
A fewmodificationsof this methodmake itpossibletoprepare PCR-qualityfishfillet genomicDNA with
a brief incubationinhotsodium hydroxide andpHadjustmentwitha Trissolution(HotSHOT).The Hot-
SHOT methodisrapid,inexpensive andmaybe carriedoutin 96-well plates,makingitamenable to
automationandhigh-throughputgenotyping. The reagentsforHot SHOT DNA preparationare simple to
prepare.An alkaline lysisreagentwith25mM NaOH, 0.2 mMdisodiumEDTA anda pH of 12 isprepared
by dissolvingthe saltsinwaterwithoutadjustingthe pH. A neutralizingreagentwith40mM Tris-HCl and
a pH of 5 ispreparedby dissolvingTris-HCl(notTrisbase) in waterwithoutadjustingthe pH.Tissue
samplesare collectedintoa96-well thermal cyclerplate orthermal cyclerstriptubes.The tissue sample
mustbe small;toolarge a sample can cause the methodto fail.Alkaline lysis reagent(75mLisaddedto
the samples andheatedto95°C for 10 minto 1 h. The undissolvedtissuedoesnotinterfere withPCR.
Afterheating,samples are cooledto4°C, and 75mL neutralizing reagentare addedtoeachsample. One
to five microlitersof the final preparationare usedpereach10-mLPCR volume.The combinationof the
alkaline lysisandneutralizingreagents yieldsabufferconsistingof 20 mM Tris-HCl (pH8.1) and 0.1 mM
EDTA, whichissimilartoa common DNA storage buffer.
The DNA extractionprocessrequirescareful handlingof biological materialtopreventsample
contaminationandcrossover.Tubesshouldbe carefullylabeled,especiallywhentransfersare required.
Robotsmay be employedtoextractreference samplesandsome evidence samples,butother
evidentiarysamplesmayrequire the directattentionof aDNA analyst.