Comparative Proteomics Kit I:
Protein Profiler Module – Quick
Guide
Lesson 1 Quick Guide
1 Label one 1.5 ml fliptop micro tube for each of
five fish samples. Also label one screwcap micro
tube for each fish sample.
2. Add 250 µl of Bio-Rad Laemmli sample buffer to
each labeled fliptop microtube.
3. Cut a piece of each fish muscle about 0.25 x 0.25
x 0.25 cm
3
( ) and transfer each piece into a
labeled fliptop micro test tube. Close the lids.
4. Flick the microtubes 15 times to agitate the tissue
in the sample buffer.
5. Incubate for 5 minutes at room temperature.
6. Carefully transfer the buffer by pouring from each
fliptop microtube into a labeled screwcap
microtube. Do not transfer the fish!
7. Heat the fish samples in screwcap microtubes
for 5 minutes at 95°C.
Lesson 2 Quick Guide
1. Set up Mini-PROTEAN 3 gel box and add 1x TGS
electrophoresis buffer to the chamber.
2. Prepare a Ready Gel cassette by cutting
along the black line on the bottom of the
cassette with a razor blade and pulling off
the plastic strip, as indicated on gel
cassette.
3. Remove the comb from the Ready Gel
cassette.
4. Place Ready Gel cassette into the
electrode assembly with the short plate
facing inward. Place a buffer dam or
another Ready Gel cassette on the
opposite side of the electrode assembly,
with notch on buffer dam facing inward.
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QUICK GUIDE
5. Slide gel cassette, buffer dam, and
electrode assembly into the clamping
frame.
6. Press down the electrode assembly while
closing the two cam levers of the
clamping frame.
7. Lower the inner chamber into the mini
tank.
8. Completely fill the inner chamber with 1x
TGS electrophoresis buffer, making sure
the buffer covers the short plate (~150 ml).
9. Fill mini tank approximately 200 ml of
1x TGS electrophoresis buffer.
10. Plate sample loading guide on top of the
electrode assembly.
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Student Manual
Sample
Loading
Guide
Pipet Tip
QUICK GUIDE
11. Heat fish samples and actin and myosin
standard to 95°C for 2–5 min.
12. Load your gel:
Lane Volume Sample
1 & 2 empty empty
3 5 µl Precision Plus Protein Kaleidoscope
prestained standards (Stds)
4 10 µl fish sample 1
5 10 µl fish sample 2
6 10 µl fish sample 3
7 10 µl fish sample 4
8 10 µl fish sample 5
9 10 µl actin and myosin standard
(AM)
10 empty empty
13. Electrophorese for 30 minutes at 200 V in 1x
TGS electrophoresis buffer.
14. After electrophoresis, remove gel from cassette
and transfer gel to a container with 25 ml Bio-Safe
Coomassie blue stain per gel and stain gel for
1 hour, with gentle shaking for best results.
Lesson 3 Quick Guide
1. Discard stain and destain gels in a large
volume of water for at least 30 minutes to overnight,
changing the water at least once. Blue-stained
bands will be visible on a clear gel after destaining.
2. Dry gels using GelAir cellophane.
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Student Manual
Banana Plugs
Anode Banana Plug
(Red)
Notch on
U-Shaped Gasket
Gel Sandwich
Pressure Plate
Cams
Mini Tank
Inner
Chamber
Assembly
Clamping
Frame
Electrode
Assembly
Cathode Banana Plug
(Black)
Lid
Fig. 11. Assembling the Mini-PROTEAN 3 cell.
QUICK GUIDE
Bio-Rad
Laboratories, Inc.
Life Science
Group
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