Bos Taurus Tissue Essay
1139 WordsNov 17, 20145 Pages
Lab 2-3 Lab Report
Analysis of Protein, Carbohydrates, and Triglycerides in Bos taurus Tissue
I) Materials and Methods:
Homogenates were provided made from liver, kidney, or heart in a 1:20 ratio with sucrose-phosphate buffer that was stored at -70° C. The tissue I tested was the liver homogenate. To qualitatively analyze proteins of the homogenate, 5µl of original liver homogenate was combined with 2 µl of protein gel sample buffer and heated in a water bath at 80° C for 10 minutes. After centrifuging the sample for 5 seconds, the contents were loaded onto a polyacrylamide protein gel set at 100V for Electrophoresis and stained Coomassie blue for an hour and results were observed the next class period a week later (Clendening).
To…show more content… Glycerol standard sample was given to the class. One sample of 3 ml Triglyceride reagent was heated at 37°C for 5 minutes, then mixed with 30 µl of Bos taurus homogenate and incubated for 10 more minutes at the same temperature. Absorbance of the glycerol standard and homogenates were measured, and converted to concentration of glycerol (Clendening).
( A520 homogenate / A520 of the standard ) x 2.5 mg/ml glycerol
The average triglyceride amount for each homogenate was determined.
To make a standard curve of glucose, 6 samples of 3 ml of Trinder reagent were heated at 37°C. After 5 minutes, 30 µl of 0.2 M citrate buffer was added to one sample, 30 µl of 0.2, 0.4, 0.6, 0.8, and 1.0 were added to one of the remaining 5 samples of Trinder reagent and heated for 10 minutes at the same temperature. Absorbance of the standards was measured (Clendening).
Y= .3985x - .0005, R2 = .9871 To measure the free glucose and glycogen, 2 samples of 3 ml of Trinder reagent were warmed at 37°C. After 5 minutes, from the digestion of glycogen process, 30 µl of the amyloglucosidase-treated reaction was added to one sample and 30 µl of the free glucose control was added to the other sample. The samples were heated for 10 minutes at 37°C and absorbance was recorded. Concentration of free glucose was calculated when solving for x using the absorbance of the reaction without amyloglucosidase