Syakalima M, Takiguchi M, Yasuda J, Hashimoto A
Department of Veterinary Clinical Science, Graduate School of Veterinary Medicine, Hokkaido University, Japan.
Zentralbl Veterinarmed A. 1997 Dec;44(9-10):603-10. doi: 10.1111/j.1439-0442.1997.tb01146.x.
Quantifying alkaline phosphatase (ALP) isoenzymes in canine serum would provide a useful index in a clinical laboratory. To achieve this goal, we tested a semi-automatic assay combining wheat germ lectin (WGL) precipitation and chemical inhibition of isoenzymes of the TNS gene with levamisole to quantify bone ALP (BALP) and corticosteroid-induced ALP (CALP), respectively. The liver ALP (LALP) isoenzyme was then calculated from the equation: TALP = BALP + LALP + CALP BALP, LALP and CALP standards from serum of puppies, bile-duct ligated dogs and dogs on 4.4 mg/kg/day prednisolone for 30 days, respectively, were used. The suitability of standard sera was tested by affinity electrophoresis. Levamisole (4.2 mM) inhibits 98% of BALP and LALP but only 42% CALP. Multiplying measured CALP by 1.8 gives the total CALP value in serum. WGL precipitated 92.3% BALP, 23.3% LALP and 26.8% heated CALP standards. These values were used to adjust precipitated ALP to obtain the exact levels of BALP. WGL was then tested on pooled serum standards in which the relative proportions of all the ALPs were known and controlled. BALP was adequately quantified except when LALP and CALP levels were extremely high. The assay was also applicable under conditions resulting in high ALP. Therefore, combining WGL and levamisole inhibition provides an adequate separation and quantification of canine ALP isoenzymes. The method has great potential for diagnostic use and should be tested further for routine implementation.
对犬血清中的碱性磷酸酶（ALP）同工酶进行定量分析，可为临床实验室提供一个有用的指标。为实现这一目标，我们测试了一种半自动检测方法，该方法结合了麦胚凝集素（WGL）沉淀法以及用左旋咪唑对TNS基因的同工酶进行化学抑制，分别用于定量骨ALP（BALP）和皮质类固醇诱导的ALP（CALP）。然后根据公式计算肝ALP（LALP）同工酶：总ALP（TALP）=BALP+LALP+CALP。分别使用幼犬血清、胆管结扎犬血清以及给予4.4mg/kg/天泼尼松龙30天的犬血清作为BALP、LALP和CALP的标准品。通过亲和电泳检测标准血清的适用性。左旋咪唑（4.2mM）可抑制98%的BALP和LALP，但仅抑制42%的CALP。将测得的CALP乘以1.8可得出血清中总CALP值。WGL沉淀了92.3%的BALP、23.3%的LALP和26.8%的热变性CALP标准品。这些值用于调整沉淀的ALP，以获得BALP的准确水平。然后在已知并控制所有ALP相对比例的混合血清标准品上测试WGL。除了LALP和CALP水平极高的情况外，BALP得到了充分定量。该检测方法在导致ALP升高的条件下也适用。因此，结合WGL和左旋咪唑抑制作用可对犬ALP同工酶进行充分的分离和定量。该方法具有很大的诊断应用潜力，应进一步进行测试以用于常规检测。
