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TECH NOTES:
Catalase activity assay based on an azo dye
Chemiluminescence detection
Detection of reactive oxygen species
Measurement of L-lactate in biological samples
Streamlined plasmid DNA purification
Recombinant adenovirus for gene expression
Differential detection of multiple DNA-binding complexes
Tetrazolium salts in clinical biochemistry
Exploration of luciferase-based light reactions
Protein assay: principle and application
Catalase activity assay based on an azo dye
Chemiluminescence detection
Detection of reactive oxygen species
Measurement of L-lactate in biological samples
Streamlined plasmid DNA purification
Recombinant adenovirus for gene expression
Differential detection of multiple DNA-binding complexes
Tetrazolium salts in clinical biochemistry
Exploration of luciferase-based light reactions
Protein assay: principle and application
Biomedical Research Service
& Clinical Application
& Clinical Application
USEFUL LABORATORY RECIPES:
TE: 10 mM Tris-HCl (pH 8.0) and 1mM EDTA
100% TCA: Add 227 ml dH2O to a bottle containing 500 g TCA. Stir to dissolve TCA and store the solution at 4ºC. Note: TCA is highly corrosive.
Use extreme caution in handling the solution. Avoid skin contact and inhalation.
20x Phosphate-Buffered Saline (PBS): Stir to dissolve 8 g KCl, 320 g NaCl, 8 g KH2PO4 and 46 g Na2HPO4 in enough dH2O. The final volume
should be 2000 ml. Filter solution through a 0.4-micron membrane. Store at room temperature. Dilute the stock 20 fold with ice-cold dH2O for
cell/tissue rinsing and harvesting.
4% Paraformaldehyde: Add 450 ml dH2O to a 2-liter beaker. Then add 2.5 ml 10N NaOH to the beaker while stirring. Wearing a mask, carefully
add 20 g paraformaldehyde to the beaker and stir ~20 min at room temperature to completely dissolve paraformaldehyde. Add 25 ml 20x PBS to
the beaker followed by adding ~2 ml 10N HCl to neutralize the solution. Use pH paper strips to confirm solution pH neutrality. Adjust pH to 7 – 7.4 if
necessary. Filter solution through a 0.4-micron membrane. Store solution refrigerated. Note: Paraformaldehyde is a biohazard. Avoid skin contact
and inhalation. Discard PFA in a designated waste bottle. Do not pour used solution down the drain.
10x SDS-PAGE Running Buffer: Stir to dissolve 75 g Tris base, 360 g Glycine and 25 g SDS in enough dH2O. The final volume should be 2500
ml. Store at room temperature. Dilute the stock 10 fold with dH2O for use as running buffer in SDS-PAGE.
Coomassie Blue Staining Solution: Mix 1.5 g Coomassie Blue R-250, 50 ml Acetic Acid, 250 ml Methanol and 200 ml dH2O. Stir to dissolve
Coomassie Blue and filter solution through a Whatman #1 paper. Store at room temperature. The solution should be reused after gel staining.
Coomassie Blue Destaining Solution: Mix 250 ml Acetic Acid, 250 ml Methanol and 2000 ml dH2O. Store at room temperature. Used destaining
solution can be refreshed by placing paper towel in the solution with agitation. This protocol removes blue dye from the solution.
10x Western Transfer Solution: Stir to dissolve 75 g Tris base and 360 g Glycine in 2000 ml dH2O first. Bring the volume to 2500 ml upon
complete dissolution of Tris and Glycine. Store the solution at room temperature. To make 2000 ml working transfer solution, mix 1600 ml cold
dH2O, 200 ml 10x Western Transfer Solution and 200 ml Methanol. The working solution should be stored at 4ºC.
10x TBE: Mix 108 g Tris base, 55 g boric acid, 40 ml of 0.5M EDTA (pH 8.0) and dH2O in a final volume of 1000 ml. Stir to completely dissolve
contents. Store at room temperature. Dilute the stock 10 fold with dH2O for gel electrophoresis.
50x TAE: Dissolve 242g Tris base and 37.2 g Na2•EDTA•(2H2O) in 900ml of dH2O first. Add 57 ml of Acetic Acid and adjust the final volume with
dH2O to 1000 ml. Store at room temperature.
0.5M EDTA: Add 186.1 g disodium ethylenediamine tetraacetate•2H2O (EDTA) to 800 ml dH2O. Stir solution vigorously while adjusting the pH to
8.0 with NaOH. EDTA will slowly go into solution as the pH approaches 8.0. Filter solution through a 0.4-micron membrane. Store at room
temperature.
LB: Stir to dissolve 10 g tryptone, 5 g yeast extract and 5 g NaCl in 1000 ml dH2O. Autoclave to sterilize. Store at room temperature.
SOC: Add 2 g tryptone, 0.5 g yeast extract, 1 ml of 1M NaCl and 0.25 ml of 1M KCl to 97 ml of dH2O. Stir to completely dissolve contents.
Autoclave to sterilize and cool to room temperature. Add 1 ml of sterile 2M magnesium mix solution (1M MgCl2 and 1M MgSO4) and 1 ml of sterile
2M glucose. Store at room temperature.
TE: 10 mM Tris-HCl (pH 8.0) and 1mM EDTA
100% TCA: Add 227 ml dH2O to a bottle containing 500 g TCA. Stir to dissolve TCA and store the solution at 4ºC. Note: TCA is highly corrosive.
Use extreme caution in handling the solution. Avoid skin contact and inhalation.
20x Phosphate-Buffered Saline (PBS): Stir to dissolve 8 g KCl, 320 g NaCl, 8 g KH2PO4 and 46 g Na2HPO4 in enough dH2O. The final volume
should be 2000 ml. Filter solution through a 0.4-micron membrane. Store at room temperature. Dilute the stock 20 fold with ice-cold dH2O for
cell/tissue rinsing and harvesting.
4% Paraformaldehyde: Add 450 ml dH2O to a 2-liter beaker. Then add 2.5 ml 10N NaOH to the beaker while stirring. Wearing a mask, carefully
add 20 g paraformaldehyde to the beaker and stir ~20 min at room temperature to completely dissolve paraformaldehyde. Add 25 ml 20x PBS to
the beaker followed by adding ~2 ml 10N HCl to neutralize the solution. Use pH paper strips to confirm solution pH neutrality. Adjust pH to 7 – 7.4 if
necessary. Filter solution through a 0.4-micron membrane. Store solution refrigerated. Note: Paraformaldehyde is a biohazard. Avoid skin contact
and inhalation. Discard PFA in a designated waste bottle. Do not pour used solution down the drain.
10x SDS-PAGE Running Buffer: Stir to dissolve 75 g Tris base, 360 g Glycine and 25 g SDS in enough dH2O. The final volume should be 2500
ml. Store at room temperature. Dilute the stock 10 fold with dH2O for use as running buffer in SDS-PAGE.
Coomassie Blue Staining Solution: Mix 1.5 g Coomassie Blue R-250, 50 ml Acetic Acid, 250 ml Methanol and 200 ml dH2O. Stir to dissolve
Coomassie Blue and filter solution through a Whatman #1 paper. Store at room temperature. The solution should be reused after gel staining.
Coomassie Blue Destaining Solution: Mix 250 ml Acetic Acid, 250 ml Methanol and 2000 ml dH2O. Store at room temperature. Used destaining
solution can be refreshed by placing paper towel in the solution with agitation. This protocol removes blue dye from the solution.
10x Western Transfer Solution: Stir to dissolve 75 g Tris base and 360 g Glycine in 2000 ml dH2O first. Bring the volume to 2500 ml upon
complete dissolution of Tris and Glycine. Store the solution at room temperature. To make 2000 ml working transfer solution, mix 1600 ml cold
dH2O, 200 ml 10x Western Transfer Solution and 200 ml Methanol. The working solution should be stored at 4ºC.
10x TBE: Mix 108 g Tris base, 55 g boric acid, 40 ml of 0.5M EDTA (pH 8.0) and dH2O in a final volume of 1000 ml. Stir to completely dissolve
contents. Store at room temperature. Dilute the stock 10 fold with dH2O for gel electrophoresis.
50x TAE: Dissolve 242g Tris base and 37.2 g Na2•EDTA•(2H2O) in 900ml of dH2O first. Add 57 ml of Acetic Acid and adjust the final volume with
dH2O to 1000 ml. Store at room temperature.
0.5M EDTA: Add 186.1 g disodium ethylenediamine tetraacetate•2H2O (EDTA) to 800 ml dH2O. Stir solution vigorously while adjusting the pH to
8.0 with NaOH. EDTA will slowly go into solution as the pH approaches 8.0. Filter solution through a 0.4-micron membrane. Store at room
temperature.
LB: Stir to dissolve 10 g tryptone, 5 g yeast extract and 5 g NaCl in 1000 ml dH2O. Autoclave to sterilize. Store at room temperature.
SOC: Add 2 g tryptone, 0.5 g yeast extract, 1 ml of 1M NaCl and 0.25 ml of 1M KCl to 97 ml of dH2O. Stir to completely dissolve contents.
Autoclave to sterilize and cool to room temperature. Add 1 ml of sterile 2M magnesium mix solution (1M MgCl2 and 1M MgSO4) and 1 ml of sterile
2M glucose. Store at room temperature.