Conti M, Gelfi C, Righetti P G
Faculty of Sciences, Department of Cell Biology, University of Calabria in Arcavacata di Rende Cosenza, Italy.
Electrophoresis. 1995 Aug;16(8):1485-91. doi: 10.1002/elps.11501601246.
Separation and quantitation of the three main hemoglobin components of umbilical cord blood (fetal, acetylated fetal and adult hemoglobins; Hb F, Fac, A) by capillary isoelectric focusing (IEF) in a pH 6-8 gradient is reported. Even in coated capillaries (with covalently bound chains of linear acrylamido derivatives, notably N-acryloylaminoethoxyethanol), no base line separation is obtained between Hb F and A, although this is routinely achieved in gel slab IEF. However, when the carrier ampholyte buffers were added to 3% short-chain liquid linear polyacrylamide, base line resolution and stabilization of peak transit times were obtained. This suggests that even in the best coating procedures, patches of the inner capillary surface could still be naked, so that the static coating is complemented by a dynamic coating on the unoccupied sites. An additional improvement in separation occurs if the above mixture, comprising 5% carrier ampholytes in the pH 6-8 range and 3% soluble polyacrylamide, is made to contain 50 mM beta-alanine, a "separator" known to flatten the pH gradient around pH 7. In the normal newborns analyzed (n = 30), the following average values were obtained: Hb F, 70.1% (range 65-75%); Hb A, 20.2% (range 15-25%); and HbFac, 9.5% (range 7-11%).
报道了通过在pH 6 - 8梯度下的毛细管等电聚焦（IEF）对脐带血的三种主要血红蛋白成分（胎儿血红蛋白、乙酰化胎儿血红蛋白和成人血红蛋白；Hb F、Fac、A）进行分离和定量。即使在涂覆的毛细管中（具有共价结合的线性丙烯酰胺衍生物链，特别是N - 丙烯酰基氨基乙氧基乙醇），Hb F和A之间也无法实现基线分离，尽管在凝胶板IEF中通常可以实现。然而，当将载体两性电解质缓冲液添加到3%的短链液体线性聚丙烯酰胺中时，获得了基线分辨率和峰迁移时间的稳定性。这表明即使在最佳的涂覆程序中，毛细管内表面的某些区域可能仍然是裸露的，因此静态涂覆由未占据位点上的动态涂覆来补充。如果上述混合物（包含pH 6 - 8范围内的5%载体两性电解质和3%可溶性聚丙烯酰胺）含有50 mM β - 丙氨酸（一种已知可使pH 7附近的pH梯度变平的“分离剂”），则分离会有进一步改善。在所分析的正常新生儿（n = 30）中，获得了以下平均值：Hb F为70.1%（范围65 - 75%）；Hb A为20.2%（范围15 - 25%）；HbFac为9.5%（范围为7 - 11%）。
