Sheet1Trial 1Tube #1234567Did bleaching occurYesNoNoYesYesNoYesTime took for bleaching (min) 18nonenone168none25Trial 2Tube #1234567Did bleaching occurYesNoNoYesYesNoYesTime took for bleaching. 19nonenone1711none28Trial 3Tube #1234567Did bleaching occurYesNoNoYesYesNoYesTime took for bleaching. 16nonenone1710none27 Running Head: Lab Report Lab Report 13 Name: Erricul Harris Course : Date: To demonstrate which part of the fly thorax cell homogenate and carries out glycosis and which part carries out respiration Table of Contents 2Abstract 2Introduction 2Cell Fractionation 3Methods 3Part A: Preparation of homogenate 6Part B: Biochemical analysis of fractionated homogenate 7Results and Analysis 11Discussion and conclusion 13References Abstract This experiment is performed to demonstrate which part of the fly thorax cell homogenate and carries out glycolysis and which part carries out respiration. Flies mainly have three body parts namely the head, thorax and abdomen. The thorax is the part holding the wings and legs. The lab experiment was conducted basing the general knowledge on flies and their body parts. More importantly, it was based on good knowledge of living cells of the flies which is useful in discussing the transportation of glycolysis and carrying out of respiration in the body of a fly. The lab results were then obtained by collecting the flies whose body parts were used to test the assumption that the mitochondrion cells are found in the thorax . Color observations were recorded to determine the usage of oxygen and therefore giving required results . It was observed that glucose was used fast in the test tubes used for the experiment . It was therefore concluded that respiration takes place in the thorax of flies. This experiment is significant in finding out the location of cells used for respiration in flies. Introduction Cell Fractionation There are two major pieces of evidence that give us the details and the process on intercellular booth ; (1) evidence obtained from biological and physical separation of intercellular constituents actual (2) inferences obtained through observation by an aid of a microscope from microscopic observation(Keeton, n.d .). The experiment aims at provision of evidence for localization of the cellular respiration of glycolysis and that of mitochondria as cytoplasm soluble; hence, verify the various function of particular cell par t. This experiment furthermor e aims at a separation of different cell organelles to obtain mitochondria using sophisticated lab research to exploit on the procedure to execute these results . I chose a system that yields a high harvest of mitochondria that are keys our procedure yields an abundant harvest of mitochondria. In the experiment, I employed two methods on my research: Centrifugation and homogenization. Homo.

1. Sheet1Trial 1Tube #1234567Did bleaching
occurYesNoNoYesYesNoYesTime took for bleaching (min)
18nonenone168none25Trial 2Tube #1234567Did bleaching
occurYesNoNoYesYesNoYesTime took for bleaching.
19nonenone1711none28Trial 3Tube #1234567Did bleaching
occurYesNoNoYesYesNoYesTime took for bleaching.
16nonenone1710none27
Running Head: Lab Report
Lab Report
13
Name: Erricul Harris
Course
:
Date:
To demonstrate which part of the fly thorax cell homogenate
and carries out glycosis and which part carries out respiration
Table of Contents
2Abstract
2Introduction
2Cell Fractionation
3Methods
3Part A: Preparation of homogenate
6Part B: Biochemical analysis of fractionated homogenate
7Results and Analysis
11Discussion and conclusion
13References
Abstract

2. This
experiment is performed to demonstrate which part of the fly
thorax cell homogenate and carries out glycolysis and which
part carries out respiration. Flies mainly have three body parts
namely the head, thorax and abdomen.
The thorax is the part holding the wings and legs. The lab
experiment was conducted basing the general knowledge on
flies and their body parts.
More importantly, it was based on good knowledge of living
cells of the flies which is useful in discussing the transportation
of glycolysis and carrying out of respiration in the body of a
fly. The lab results were then obtained by collecting the flies
whose body parts were used to test the assumption that the
mitochondrion cells are found in the thorax
. Color observations were recorded to determine the usage of
oxygen and therefore giving required results
. It was observed that glucose was used fast in the test tubes
used for the experiment
. It was therefore concluded that respiration takes place in the
thorax of flies. This experiment is significant in finding out the
location of cells used for respiration in flies.
Introduction
Cell Fractionation
There
are two major pieces of evidence that give us the details and
the process on
intercellular booth
; (1) evidence obtained from biological and physical separation
of intercellular constituents actual (2) inferences obtained
through observation by an aid of a microscope from microscopic
observation(Keeton, n.d
.). The experiment aims at provision of evidence for localization
of the cellular respiration of glycolysis and that of mitochondria

3. as cytoplasm soluble; hence, verify the various function of
particular cell par
t. This experiment furthermor
e aims at a separation of different cell organelles to obtain
mitochondria using sophisticated lab research to exploit on the
procedure to execute these results
. I
chose a system that yields a high harvest of mitochondria that
are keys our procedure yields an abundant harvest of
mitochondria.
In the experiment, I employed
two methods on my research: Centrifugation and
homogenization. Homogenization produces a solution is which
consist of a suspension which is insoluble and soluble cell
constituents.
The suspension contains intracellular as reference results which
have mitochondria organelles
(Raven & Johnson, n.d.). Suspension and soluble cell
components were separated using centrifugal.
The material was spun in a centrifuge machine on its axis
where centrifugal force caused the heavy stuff (suspension)
directed the axis outwards. The suspension moved and collected
at the end of the centrifuge tube.
My choice of experimental material
is the need to obtain a high number of respiratory and glycolytic
and separability of mitochondria from the soluble hence my
selection of insect flight muscle. As for my hypotheses, the null
hypothesis (H0) is that there is an increase of glycolysis and
respiration enzymes in cells, whereas the alternate hypothesis
(H1) is that there is no significant respiration process relation
glycolysis and respiration in insect wing
.Methods
Part A: Preparation of homogenate

4. The instructor assigned one team of two students to prepare
homogenizer.
The team was to obtain a clean homogenizer and chilled it for
5-10 minutes before use in an ice bath
. The instructor distributed 60 flies among the other teams as
the one team was obtaining the homogenizer. The flies were
immobilized by keeping them in a closed plastic tube on ice
because cold causes anesthesia in insects. The wings, legs,
heads, and abdomens were cut off quickly by razor blades from
every fly. The cut was done quickly to avoid warming
. The thorax of each fly was then saved. The entire class did the
cutting simultaneously as fast as possible.
Each team
was to cut each thorax in half to facilitate grinding tissue. The
thoraces were then put into chilled glass homogenizer tube as it
sat on ice(Biology 155 Laboratory Supplement
, n.d.).
15.0 ml of ice-cold homogenizing medium was added to the
homogenizer tube i.e. 0.32M mannitol containing 0.02M
phosphate buffer, pH 7.4.
The team that was designated to prepare homogenizer made the
homogenate for the entire class.
The homogenizer was then run up and down into the mix of
medium and thoraces until the mixture became thick (like a
milkshake). During the process, the homogenizing tube was kept
on the ice. A different pair of students, assigned before,
prepared a filtering device which consisted of a 5inch diameter
circle of cheesecloth, two layers thick, wetted with
homogenizing medium (but not dripping), placed in a short-
stemmed glass funnel. The center of the cheesecloth was pushed
down as far as the beginning of the stem. The stem, in turn, was
fitted into a 50ml graduated cylinder. It helped to put the
cylinder in a beaker of ice.

5. The homogenate was then transferred to the cheesecloth where
it began to filter through. The funnel was to be slightly shaken
if the filtration did not start immediately.
For the case where the shaking could not still work, it was
squeezed through by clean hand.
An additional 10ml of the ice-cold medium was added to the
homogenizing vessel and the homogenizer run to suspend any
residual tissue debris. The medium was then transferred to the
cheesecloth in the funnel and was used to wash down the
material trapped on the cheesecloth. The washing of the
homogenizer and the cheesecloth was repeated with an
additional 5.0ml of ice-cold medium. The cheesecloth bag was
finally squeezed into the funnel with clean hands. The filtration
provided 30ml of the homogenate in the cylinder while the
materials hold back in the cheesecloth were mostly pieces of
thoracic integument and large muscle fibers.
Thoroughly filtered homogenate was then mixed (the mixing is
crucial), and the total volume was recorded to the nearest
milliliter. 15ml of the homogenate was then transferred to a
clean tube which was marked H (for the whole homogenate).
The tube was kept on the ice every time
. The remaining 15ml homogenate was transferred to a clean
centrifuge tube placed in a beaker of crushed ice. A balance
tube was prepared by putting 15ml distilled water into a new
centrifuge tube like the one used before
. Both tubes were then placed in the refrigerated centrifuge, on
opposite sides of the rotor and centrifugation was done at
5000rpm for 20 minutes(Biology 155 Laboratory Supplement,
n.d.).
Another crucial step was to stop the centrifuge immediately,
and the tube containing the homogenate was retrieved, carefully
holding it at the same angle at which it laid in the centrifuge.
All the supernatant was poured into a clean (rinsed with

6. distilled water and shaken dry) 25ml graduated cylind
er. The pellet (whitish in color) was not poured to achieve a
"clean"
separation. The volume of the supernatant was restored to 15ml
with the homogenizing medium, shaken well to mix the
contents. The contents were then transferred to a clean tube,
marked S, and kept on ice. The volume of S was exactly equal
remaining 15ml homogenate that had been moved to clean
centrifuge before
.
The ice-cold homogenizing medium was then added to the
pellet. The amount of medium added was made to be just
enough to make the final volume of re-suspended pellet exactly
equal to the original volume centrifuged (15.0ml) (This step
was also crucial).
The tube was the stoppered and shaken to resuspend the pellet
thoroughly. When the pellet was re-suspended, it was labeled P.
The three labeled tubes for un-centrifuged homogenate (H), re-
suspended pellet (P) and supernatant (S) were then place on ice.
The pellet contains nuclei, glycogen (polysaccharide) granules,
mitochondria, and bits of the muscle's contractile apparatus.
The supernatant contains most soluble muscle constituents,
including glycolytic enzymes and some membranous material
(reticulum) which is too small to centrifuge out at the speeds
used. Finally, 1ml of H, 1ml of S, and 1ml of P was then
obtained in the labeled tubes and kept in a beaker of crushed ice
Part B: Biochemical analysis of fractionated homogenate
Each team of two students obtained the following materials in
the quantities indicated. They were kept in clean, dry, labeled
test tubes in a rack at room temperature and not on the ice.
Substance
Concentration
Quantity

7. Mannitol buffer*
0.32M
3ml
Buffer-cofactor-dye mixture**
5ml
Glucose
0.015M
3ml
Succinate
0.2M
3ml
Table 1
* Mannitol buffer = homogenizing medium
** Potassium phosphate buffer, pH 7.4 (0.2M); ATP (0.0125M);
MgCl2 (0.005M); Methylene blue (0.5mg/ml).
1ml plastic pipettes for these solutions and three pipettes for
dealing with tubes H, S and P were obtained. Each pipette was
marked (to avoid confusion) and left in the tube
. Seven clean, small glass test tubes (identical in size and
numbered 1-7 using a grease pencil) were used as reaction
vessels. A pan containing water adjusted to 35 degrees Celsius
and about two inches deep was also used
(Biology 155 Laboratory Supplement, n.d.).
The table below shows what was added to each of the seven
reaction tubes:
Ingredient/Reaction Tube Number
1
2
3
4

8. 5
6
7
Mannitol
0.25
0.25
0.25
0.25
0.25
0.25
0.25
Buffer mix
0.45
0.45
0.45
0.45
0.45
0.45
0.45
Glucose
0.20
0.20
0.20
0.20
0
0
0
Succinate
0
0
0
0
0.20
0
0
Whole Homogenate (H)

9. 0
0
0.25
0.25
0
0
0.25
Pellet (P)
0
0.25
0
0.25
0.25
0
0
Supernatant
0
0
0.25
0.25
0
0.25
0
Table 2
The rack of reaction tubes was placed in the pan of water at 35
degrees Celsius and time were recorded. The temperature was
made constant at 35 degrees Celsius. The exact time at which
individual tubes lost blue color was recorded.
Results and Analysis
Results

10. Trial 1
Tube #
1
2
3
4
5
6
7
Did bleaching occur
Yes
No
No
Yes
Yes
No
Yes
Time took for bleaching (min)
18
None
none
16
8
none
25

11. Trial 2
Tube #
1
2
3
4
5
6
7
Did bleaching occur
Yes
No
No
Yes
Yes
No
Yes
Time took for bleaching.
19
None
none

12. 17
11
none
28
Trial 3
Tube #
1
2
3
4
5
6
7
Did bleaching occur
Yes
No
No
Yes
Yes
No
Yes

13. Time took for bleaching.
16
None
none
17
10
none
27
Table 3.Table showing Trials on test tubes
The ingredients used include the following:
Homogenized solution of mannitol at a concentration of 0.320M
3.0ml, Succinate solution of 0.20M 3.0ml, 5.0 ml homogenized
cofactor of dye mixture, and 3.0ml of 0.015 M glucose
.
Fig 1.1 Test Trial 1
Fig1.2 Trial 2
Fig 1.3 Trial 3
From the results, there is no color change in tests tube numbers
two, three, and six
In test tube four, the time taken is seventeen minutes for trial 1
and two while trial one took sixteen minutes for the same
number of test tube.
In all the test tubes, number seven (7) to the longest time in all
the trials with trial 2 being the highest at twenty eight
.

14. Discussion and conclusion
Consider the reagents table below;
Concentration of Ingredients
Ingredients/ Test tube number
1
2
3
4
5
6
7
Mannitol
0.25
0.25
0.25
0.25
0.25
0.25
0.25
Buffer mix
0.45
0.45
0.45
0.45
0.45
0.45
0.45

15. Glucose
0.2
0.2
0.2
0.2
0
0
0
Succinate
0
0
0
0
0.2
0
0
Whole Homogenate
0
0
0.25
0.25
0
0
0.25
pellet (P)
0
0.25
0
0.25
0.25
0
0
Supernatant
0
0
0.25

16. 0.25
0
0.25
0
Figure 2. Table showing concentration of fractionated
homogenate used
The water was kept at a level of the that makes the test tubes
slightly be it and maintained at a constant temperature of 35.0
Celsius and reason for this was to allow the reagents to be
active, hence the reason for adding hot water to the water bath
As noted earlier, the indicator of reaction in the tube is dye
color. The color of methylene
changed from blue to colorless and was seen from test tubes 1,
4, 5, and 7.
Test tube 1 and test tube 7 has the same components except that
test tube one has glucose and seven has whole Homogenate
, The absence of glucose in test tube seven resulted to test tube
seven taking longer time than any test tube in a color change for
all the trials. This is a great analogy that we obtained meaning
that glucose was used fast in the test tubes as the cells use it to
produce energy through glycolysis and respiration resulting in
changing of color(Baker & Allen, n.d.). Also, glucose is the
initial substrate of respiration its process and glycolysis. Our
discussion also sets that we fail to reject our hypothesis, Ho,
that there is an increase of glycolysis and respiration enzymes
in cells at presence of cells containing mitochondria. Flight
muscle contains a great number of mitochondria, hence, cells
from the centrifugal process yielded our desired results(Loewy
& Siekevitz, n.d.)
References
Baker, & Allen,. The Study of Biology (3rd ed.). 195-232.

17. Biology 155 Laboratory Supplement.
Keeton,. Biological Sciences (3rd ed., pp. 137-138,163-180).
Loewy, & Siekevitz,. Cell structure and Function (2nd ed., pp.
310-314).
Raven, & Johnson,. Biology (4th ed., pp. 193-205.).
�Take out the Running Head: Lab Report. Put your last name
with the page number.
�Put BIOL 155 lab for course and the date for date and put all
of that with your name below the title. You should also put
your lab partners.
�The title needs to be reworded. It should have the scientific
name of the fly, what part of the fly was used, that cell
separation was uses, and that we were looking for the cellular
location of glycolysis and respiration in the cell. The part
where you say “and carries out glycolysis and which part carries
out respiration” should be reworded. It should say something
like “subcellular site of glycolysis and respiration”. Also you
could say determination instead of “to demonstrate”.
�This part is not needed, but it is cool that you put it in.
�Indent
�This does not need to be in the abstract.

18. �
�This doesn’t need to be in the abstract
�Say action instead.
�Take out the “the lab results were then obtained” part. It
would be better to take out this sentence and put something like
mitochondria and cytoplasm were separated by……..
�Showing respiration took place.
�You sure that glucose was the fastest. Look at the results.
You should also say what tubes bleached, and why did they
bleach.
�Expand on this a little more, why would knowing this
information be important or other scientist. Relate this to
something that could be in a real life research setting.
�You need to talk about why the fly was used, what glycolysis
and respiration in depth, and what the purpose of the experiment
is. You don’t clearly say what the purpose is.
�Indent

19. �on
�of
�Processes
�Make sure that you don’t directly quote the author, you have
to paraphrase.
�You need to put the year here.
�parts
�Wordy not needet
�Take this out.
�
�Take this sentence out.
�Take this out and reword the sentence in passive voice.
�Look at what homogenization means. Also the sentence
doesn’t make sence.

20. �This sentence does not make sence.
�
�Look up what centrifugation does.
�Don’t bring yourself into the paper, and this sentence is
repetitive.
�This is not what are experiment is about. You hypothesis
should be what tubes will bleach and what ones wont and which
ones you think will be faster. Just make a normal hypothesis
instead of a null hypothesis. A null hypothesis will make this
more complicated for you.
�Take this out
�Reword this to say a clean homogenizer was chilled in a ice
bath for 5-10 minutes.
�You can add the to avoid warming to the previous sentence.
�You can take this out.
�Take out each team.

21. �Put the authors last name and the year with none of it being in
italics.
�
�Good that you put that.
�Take out
�?
�This is understood.
�Take the first part out, and reword the last part to say the
cheesechloth was squeezed by hand.
�Take this part out.
�Take this out.
�You don’t have to add info about the balance tube.
�The supernatant is a liquid it is never dry and not distilled

22. water was added.
�Say to separate mitochondria and cytoplasm instead of clean
separation.
�Take this sentence out it is not needed.
�Take this part out.
�Take this out.
�Take this whole table out.
�This part is understood and not needed.
�Why was it kept at 35C and take out also and add the answer
to this question at the end of the sentence.
�You need to reference the table in the text.
�What are the units.
�This goes on top of the table
�You don’t need this because you have this in the figure.

23. �This should be in the methods.
�There should be only one figure. You need to average all
three trials for each tube and put that into one graph. The graph
should be a bar graph. A line graph is used for examining one
treatment over time. We are analyzing the effect of 7 different
treatments (different chemicals and cell parts added to each
tube) at once.
�
�Reference the one figure you will have in your rough draft and
mention which tube took the shortest time to bleach and the
longest time to bleach (which you have already for the longest
which is good).
�You don’t need this table again.
�This should not be in the discussion.
�Methylene blue
�Dose not have glucose.
�The discussion should be the longest part of the paper. You
should explain why each tube bleached or not (explain what cell

24. parts were needed to break down the sugar), why some tubes
bleached faster than other. You should also relate this project
to research from a research article.
�Look in the lab supplement on how to write the references.