Bradford Assay

in this video you will learn how to perform protein quantitation using the Bradford assay protein quantitation is one of the most commonly performed procedures in a biotechnology laboratory and this activity the protein concentration and milk will be measured using the Bradford assay for this activity you will need 1x Bradford reagent and 1x phosphate buffered saline or PBS using a 22200 microliter micropipette set the pipette to 98 microliters pipette ninety-eight microliters of 1x PBS into an empty microcentrifuge tube labeled sample you switch to a to-220 microliter micropipette and set the pipette to 2 microliters pipette to microliters of milk sample into the micro tube labeled sample a you mix the milk sample and PBS by either pipetting or vortexing this mixture has diluted the milk to one part in 50 labeled to cuvettes one control and the other sample do not handle the cube yet where the light passes through and make certain your labels are well above this area pipette 20 microliters of diluted milk sample into the cuvette labeled sample and 20 microliters of 1x PBS into the cuvette labelled control next we will set up protein standards see your protocol for the correct concentrations label a cuvette for each standard and mark it with the corresponding concentration next add 20 microliters of each protein standard into the corresponding cuvette make certain you use a clean pipette tip for each sample you now you are ready to add the Bradford reagent to all of the cue gets you have prepared including the sample and control add one milliliter of 1x Bradford reagent to each of the cuvettes mix completely by pipetting up and down with the micropipette incubate the cuvettes at room temperature for five minutes after five minutes visually compared the cuvette containing the milk sample to the cuvettes containing the protein standards determine the standard that most closely matches the color of the milk sample estimate the protein concentration of the milk sample based upon the visual comparison to determine the protein tration using a spectrophotometer select for the Bradford assay when asked insert blank into the spectrophotometer insert the cuvette labeled control use the blank to set the spectrophotometer to zero absorbance or 100 percent transmittance if instructed read the absorbance of the seven protein standards and record the absorbance values remove the cuvette label blank and insert the milk sample into the spectrophotometer read the absorbance and record the value you

Channel: Bio-Rad Laboratories

Views: 208,841

Rating: 4.8818097 out of 5

Keywords: bio-rad, life science, education, biotechnology, biology, molecular biology, technique, method, student, teacher, school, classroom, laboratory, laboratory skills course, textbook, lab, activity, demo, demonstration, instuctional, instruction, Protein, quantitation, kit, spectrophotometer, spec, spectrophotometry, colorimetric, color, change, visual, concentration, absorbance, vis, stem, ngss

Id: vfY3mVOlGBU

Length: 5min 8sec (308 seconds)

Published: Wed Oct 17 2012