Polyclonal-antibody-based ELISA to detect milk alkaline phosphatase.
Polyclonal antibodies (PAb) prepared against bovine milk alkaline phosphatase (ALP) were used to develop a competitive indirect (CI) ELISA. Anti-ALP PAb were specific for milk ALP and did not react with ALP from E. coli or bovine and calf intestinal mucosa. Anti-ALP PAb were 20% cross-reactive with bovine placenta ALP. The anti-ALP antibodies also did not recognize bovine serum albumin, acid glycoprotein, ovalbumin, ferritin, and casein, although some cross-reactivity was observed with whey protein isolate. Anti-ALP PAbs reacted with deglycosylated native ALP, but did not recognize ALP denatured at 100 degrees C in 2% SDS or deglycosylated denatured ALP. When buffered solutions of milk ALP were heated at 70 degrees C, ALP activity decreased at a faster rate than ALP content determined by CI-ELISA. The ELISA differentiated between native and heat denatured ALP. Further studies are warranted to determine if an ELISA can be used to verify pasteurization of fluid milk.
Vega-Warner
A V
AV
Michigan State University, Department of Food Science and Human Nutrition, East Lansing 48824, USA.
Gandhi
H
H
Smith
D M
DM
Ustunol
Z
Z
D000469:Alkaline Phosphatase / Q000032:analysis*; D000818:Animals; D000906:Antibodies; D002417:Cattle; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D008892:Milk / Q000201:enzymology*; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D012680:Sensitivity and Specificity
D000906:Antibodies; D000469:Alkaline Phosphatase