Centro de Ciências Naturais e Exatas, Departamento de Química, Laboratório de Enzimologia Toxicológica, Universidade Federal de Santa Maria, Campus Universitário, Sala 2208, 97105-900, Santa Maria, RS, Brazil.
Anal Biochem. 2011 Mar 1;410(1):34-9. doi: 10.1016/j.ab.2010.10.039. Epub 2010 Nov 6.
Methods for the isolation of peripheral blood mononuclear cells (PBMCs) and human lung mononuclear cells (LMCs) have been proposed previously. This study describes a method that allows the separation of lymphocyte-rich LMCs from rats. Trypan blue was applied to determine cell viability. White blood cell and differential cell counts were also performed. Relationships between nucleoside triphosphate diphosphohydrolase (NTPDase, EC 3.6.1.5) activities expressed in milligrams of protein, millions of cells, and millions of viable cells were examined as linear correlations. The lung tissue yielded 82.46% lymphocytes, 8.6% macrophages, 2.20% monocytes, and 1.27% polymorphonuclear cells (PMNs). In LMCs, a very strong correlation was observed as follows: between NTPDase activity, as determined using ATP or ADP as a substrate, expressed in milligrams of protein and that expressed in millions of cells (r ≥ 0.91), between that expressed in milligrams of protein and that expressed in millions of viable cells (r ≥ 0.91), and between that expressed in millions of cells and that expressed in millions of viable cells (r ≥ 0.98). Based on our results, we affirm that NTPDase activity could be expressed in millions of viable cells, millions of cells, or milligrams of protein.
先前已经提出了分离外周血单核细胞（PBMCs）和人肺单核细胞（LMCs）的方法。本研究描述了一种可从大鼠中分离富含淋巴细胞的 LMCs 的方法。台盼蓝用于确定细胞活力。还进行了白细胞和差异细胞计数。研究了以毫克蛋白质、百万个细胞和百万个活细胞表示的核苷三磷酸二磷酸水解酶（NTPDase，EC 3.6.1.5）活性之间的关系，作为线性相关性进行了检查。肺组织产生 82.46%的淋巴细胞、8.6%的巨噬细胞、2.20%的单核细胞和 1.27%的多形核细胞（PMN）。在 LMCs 中，观察到以下非常强的相关性：使用 ATP 或 ADP 作为底物测定的 NTPDase 活性，以毫克蛋白质和百万个细胞表示（r≥0.91），以毫克蛋白质和百万个活细胞表示（r≥0.91），以及以百万个细胞和百万个活细胞表示（r≥0.98）。基于我们的结果，我们断言 NTPDase 活性可以以百万个活细胞、百万个细胞或毫克蛋白质表示。
