Bio-Rad Protein Assay - Manual (Page 14)

12
Section 4
Troubleshooting Guide
1. How can protein with a typical low dye response be assayed?
Occasionally a protein will be assayed which yields exceptionally low
color response to the Bio-Rad Protein Assay. One such protein is
gelatin. Although the standard range (20-140 µg) is not detectable
when the standard assay procedure is used, quantitation of the protein
is possible when the sample-to-dye ratio is changed. By using the
sample-to-dye ratio of the microassay procedure (800 µl sample + 200
µl dye reagent concentrate) a usable standard curve for moderate con-
centrations of gelatin is produced.
Therefore, for a protein which yields exceptionally low color response
to the Bio-Rad Protein Assay, quantitation in the standard range may
be possible when the microassay sample-to-dye ratio is used.
Other modifications of this dye binding assay to increase sensitivity
have been reported.
7,8
2. Absorbance of the protein solution is very low. What is the likely
cause?
The dye reagent concentrate may be old. If it is over 1 year old,
replace with a new bottle of reagent.
The sample may contain a substance which interferes with the reac-
tion, such as detergent or sodium hydroxide. Check the compatibility
guide (Table 1). Dilute the sample or switch to the DC Protein Assay
if necessary.
The molecular weight of the protein is low; the lower limit of detec-
tion for this method is approximately 3,000 to 5,000 daltons.
LIT33C 8/31/98 02:25 PM Page 12