Agarose gel electrophoresis pattern of serum alkaline phosphatase isoenzymes in Holstein cows during lactation.

A recent study found that an agarose gel electrophoresis (AGE) method yielded two distinct major bands corresponding to the hepatic and bone ALP isoenzymes (ALP2 and ALP3, respec-tively) in bovine serum treated with protease and neuraminidase (PN-treatment), although there were concerns that the intestinal ALP isoenzyme (ALP5) often overlapped with ALP3 in human serum treated with neuraminidase. Because ALP5 was separated from ALP3 in bovine serum treated with protease alone (P-treatment), we used a modified method employing both P- and PN-treated bovine sera to measure the activities of the three ALP isoenzymes in 53 lacta-ting Holstein cows: 24 primiparous and 29 multiparous. Upon electrophoresis, 51 of 53 samples (96.2%) subjected to P-treatment yielded a distinct fraction corresponding to ALP5, as did the control serum. All PN-treated sera yielded a definite ALP2 fraction. The ALP3 fraction was calculated as the remainder after excluding ALP2 and ALP5. The activities of total ALP (t-ALP) and ALP3 in primiparous cows were higher than those in multiparous cows (p ⟨ 0.001) at early-to-peak [10-110 days in milk (DIM)] and mid (111-220 DIM) lactation. In the multi-parous cows, the ALP3 activity at late lactation (221-477 DIM) was significantly higher than that at early-to-peak lactation. Thus, the modified AGE method described here is able to discrimi-nate three fractions of ALP isoenzymes in the sera of lactating cows. The AGE pattern of circu-lating ALP isoenzymes will contribute to the understanding of the physiological bone metabolism status in lactating cows.