Protein estimation in tissues containing high levels of lipid: modifications to Lowry's method of protein determination.
A method to estimate protein in detergent-solubilized homogenates of lipid-rich biological samples (e.g., adipose tissue, myelin-enriched fractions of sheep brain) is described. The method is also suitable for samples in which protein is present as a protein-detergent complex. The method involves homogenization of tissue in the presence of a suitable detergent and KCl. Protein is then estimated in an aliquot of this homogenate by Lowry's method in the presence of excess sodium dodecyl sulfate, the solutions being clarified by extraction with ethyl acetate. Protein solubilization by Triton X-100 from adipose tissue was biphasic, extracting two to three times more protein under optimum conditions [1.7 +/- 0.1% (v/v) Triton X-100 and 0.75 M KCl], compared with homogenization without salt and detergent. Unlike adipose tissue, protein solubilization from myelin-enriched fractions of sheep brain peaked at 1% (v/v) Triton X-100, resulting in the extraction of approximately three times more protein than homogenization in the absence of detergent and salt.
Upreti
G C
GC
Ruakura Agriculture Centre, Ministry of Agriculture & Fisheries, Hamilton, New Zealand.
Ratcliff
R A
RA
Riches
P C
PC
D000273:Adipose Tissue / Q000032:analysis; D000818:Animals; D001923:Brain Chemistry; D003902:Detergents / Q000494:pharmacology; D008055:Lipids / Q000032:analysis*; D009185:Myelin Proteins / Q000032:analysis; D009249:NADP / Q000378:metabolism; D011189:Potassium Chloride / Q000494:pharmacology; D011506:Proteins / Q000032:analysis*; D012756:Sheep
D003902:Detergents; D008055:Lipids; D009185:Myelin Proteins; D011506:Proteins; D009249:NADP; D011189:Potassium Chloride