Laboratorio de Patología Celular y Molecular, Centro de Medicina Experimental, Instituto Venezolano de Investigaciones Científicas, Caracas, Venezuela.
Int J Lab Hematol. 2011 Apr;33(2):194-200. doi: 10.1111/j.1751-553X.2010.01263.x. Epub 2010 Oct 7.
The IgV(H) mutational status of B-cell chronic lymphocytic leukemia (B-CLL) is of prognostic value. Expression of ZAP-70 in B-CLL is a surrogate marker for IgV(H) unmutated (UM). As determination of IgV(H) mutational status involves a methodology currently unavailable for most clinical laboratories, it is important to have available a reliable technique for ZAP-70 estimation in B-CLL. Flow cytometry (FC) is a convenient technique for this purpose. However, there is still no adequate way for data analysis, which would prevent the assignment of false positive or negative expression.
We have modified the currently most accepted technique, which uses the ratio of the mean fluorescent index (MFI) of B-CLL to T cells. The MFI for parallel antibody isotype staining is subtracted from the ZAP-70 MFI of both B-CLL and T cells. We validated this technique comparing the results obtained for ZAP-70 expression by FC with those obtained with quantitative PCR for the same patients.
We applied the technique in a series of 53 patients. With this modification, a better correlation between ZAP-70 expression and IgV(H) UM was obtained.
Thus, the MFI ratio B-CLL/T cell corrected by isotype is a reliable analysis technique to estimate ZAP-70 expression in B-CLL.
B 细胞慢性淋巴细胞白血病（B-CLL）的 IgV(H) 突变状态具有预后价值。B-CLL 中 ZAP-70 的表达是 IgV(H) 未突变（UM）的替代标志物。由于 IgV(H) 突变状态的确定涉及目前大多数临床实验室无法使用的方法学，因此拥有一种可靠的 B-CLL 中 ZAP-70 估计技术非常重要。流式细胞术（FC）是一种用于此目的的便捷技术。然而，目前仍然没有足够的数据分析方法，这可能导致假阳性或假阴性表达的错误分配。
我们修改了目前最被接受的技术，该技术使用 B-CLL 与 T 细胞的平均荧光强度（MFI）比值。从 B-CLL 和 T 细胞的 ZAP-70 MFI 中减去平行抗体同种型染色的 MFI。我们通过将 FC 获得的 ZAP-70 表达结果与同一患者的定量 PCR 获得的结果进行比较来验证该技术。
我们将该技术应用于 53 例患者。通过这种改进，我们获得了 ZAP-70 表达与 IgV(H) UM 之间更好的相关性。
因此，通过同种型校正的 B-CLL/T 细胞 MFI 比值是估计 B-CLL 中 ZAP-70 表达的可靠分析技术。
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