Purification of glycated hemoglobin free of hemoglobin A1c and its use to produce monoclonal antibodies specific for deoxyfructosyllysine [correction of deoxyfructosyllsine] residues in glycohemoglobin.
Purification of glycated hemoglobin free of hemoglobin A1c and its use to produce monoclonal antibodies specific for deoxyfructosyllysine [correction of deoxyfructosyllsine] residues in glycohemoglobin.
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纯化不含血红蛋白 A1c 的糖化血红蛋白，及其用于生产糖血红蛋白中脱氧果糖基赖氨酸 [脱氧果糖基赖氨酸的校正] 残基特异性的单克隆抗体。
DOI：
10.1016/0006-291x(91)90910-y
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发表时间：
1991
影响因子：
3.1
通讯作者：
Cohen,MP
中科院分区：
文献类型：
--
作者：
Wu,VY;Steward,LA;Cohen,MP
从人红细胞裂解物中纯化血红蛋白，并将其非酶糖化在赖氨酸残基的E-氨基上，用于BALB c小鼠的免疫。用小鼠脾细胞与SP 2 0骨髓瘤细胞融合，获得了分泌抗糖化血红蛋白单克隆抗体的杂交瘤细胞株。用纯化的单克隆抗体进行的免疫印迹显示出对糖化血红蛋白的特异性，与HbAo无反应。将红细胞裂解液应用于固定在Sepharose 4 B上的单克隆抗体亲和柱上，可有效地将糖化血红蛋白与其他血红蛋白分离。一小部分纯化的HbA 1c吸附到单克隆抗体亲和柱上，表明在同一分子中E-氨基赖氨酸和N-末端缬氨酸位置均可发生糖化。通过免疫吸附去除含有糖化赖氨酸的分子后，HbA 1c不与抗体反应，证实了抗体对脱氧果糖基-赖氨酸残基的特异性。这些结果表明，这些单克隆抗体是位点特异性的糖化赖氨酸的氨基在血红蛋白，并可以提供快速和有效的分离和鉴定人红细胞裂解物中的糖化血红蛋白。
Hemoglobin nonenzymatically glycated at E-amino groups of lysine residues was purified from human erythrocyte lysates and used for immunization of BALB c mice. Hybridomas secreting monoclonal antibodies for glycated hemoglobin were produced by fusion of mouse spleen cells with SP 2 0 myeloma cells. Immunoblotting with purified monoclonal antibody demonstrated specificity for glycated hemoglobin, with no reaction with HbA o. Glycated hemoglobin was effectively separated from other hemoglobins upon application of erythrocyte lysates to an affinity column of monoclonal antibody immobilized onto Sepharose 4B. A small fraction of purified HbA 1c adsorbed to the monoclonal antibody affinity column, indicating that glycation can occur at both E-amino lysine and N-terminal valine positions in the same molecule. HbA 1c did not react with the antibody after removal by immunoadsorption of molecules containing glycated lysine, confirming specificity of the antibody for deoxyfructosyl-lysine residues. The findings indicate that these monoclonal antibodies are site specific for glycated lysine amino groups in hemoglobin, and can provide rapid and efficient separation and identification of glycated hemoglobin in human erythrocyte lysates.