1 research outputs found
A Novel and Cost Effective Method of Removing Excess Albumin from Plasma/Serum Samples and Its Impacts on LC-MS/MS Bioanalysis of Therapeutic Proteins
We
have developed an innovative method to remove albumin from plasma/serum
samples for the LC-MS/MS quantitation of therapeutic proteins. Different
combinations of organic solvents and acids were screened for their
ability to remove albumin from plasma and serum samples. Removal efficiency
was monitored by two signature peptides (QTAÂLVEÂLVK and
LVNÂEVTÂEFÂAK) from albumin. Isopropanol with 1.0%
trichloroacetic acid was found to be the most effective combination
to remove albumin while retaining the protein of interest. Our approach
was compared with a commercial albumin depletion kit on both efficiency
of albumin removal and recovery of target proteins. We have demonstrated
that our approach can remove 95% of the total albumin in human plasma
samples while retaining close to 100% for two of three therapeutic
proteins tested, with the third one at 60–80%. The commercial
kit removed 98% of albumin but suffered at least 50% recovery loss
for all therapeutic proteins when compared to our approach. Using
BMS-C as a probe compound, the incorporation of the albumin removal
approach has improved both assay sensitivity and ruggedness, compared
to the whole plasma protein digestion approach alone. An LC-MS/MS
method was developed and validated based on this new approach for
the analysis of BMS-C in monkey serum. This assay was successfully
applied to a toxicological study. When the albumin removal method
was used in another clinical LC-MS/MS method, the sensitivity improved
10-fold to 50 ng/mL LLOQ comparing to a typical pellet digestion method