Huang Li-Chun, Lin Wendy, Yagami Machiko, Tseng Daphne, Miyashita-Lin Emily, Singh Nitasha, Lin Andy, Shih Shian-Jiun
Assay Development, Cell Genesys, Inc., 500 Forbes Blvd., South San Francisco, CA 94080, USA.
Biologicals. 2010 May;38(3):393-400. doi: 10.1016/j.biologicals.2010.01.009. Epub 2010 Feb 24.
A method using Cedex automatic cell counter (Innovatis) to determine the cell density and viability of a whole cell-based immunotherapy product has been developed and validated for the assay performance characteristics including specificity, accuracy, precision, linearity, range, and robustness. Instrument-to-instrument variation due to intrinsic differences in handmade flow cells was also evaluated. For cell density, Cedex demonstrated acceptable specificity, accuracy and precision for cell densities ranging from 3.13x10(5) to approximately 1.0x10(7)cells/mL, with intermediate precision of about 5% relative standard deviation (RSD). However, a marked difference was observed between the two instruments studied and they therefore could not be used interchangeably without additional calibration procedures that went beyond the manufacturer's recommendation. For viability, mixing known numbers of non-viable cells with highly viable cells allowed evaluation of the specificity, accuracy and linearity of the viability determination. Acceptable levels of accuracy (95.3-106.4% recovery) and precision (RSD<5%) were demonstrated for the viability range from 50 to 100%. The instrument-to-instrument difference was less than 4.6%. The assays for both cell density and viability were sufficiently robust for assay parameters. However, the effect of certain parameters was cell line-dependent, suggesting that Cedex performance should be verified for each cell line of interest.
已经开发并验证了一种使用Cedex自动细胞计数器（Innovatis公司）来测定全细胞免疫治疗产品的细胞密度和活力的方法，该方法针对包括特异性、准确性、精密度、线性、范围和稳健性等分析性能特征进行了验证。还评估了由于手工制作的流动池的内在差异导致的仪器间差异。对于细胞密度，Cedex在细胞密度范围为3.13×10⁵至约1.0×10⁷个细胞/毫升时，显示出可接受的特异性、准确性和精密度，相对标准偏差（RSD）约为5%的中等精密度。然而，在所研究的两台仪器之间观察到明显差异，因此在没有超出制造商建议的额外校准程序的情况下，它们不能互换使用。对于活力，将已知数量的非活细胞与高活力细胞混合，可以评估活力测定的特异性、准确性和线性。在活力范围为50%至100%时，显示出可接受的准确性水平（回收率为95.3 - 106.4%）和精密度（RSD < 5%）。仪器间差异小于4.6%。细胞密度和活力的测定对于分析参数而言都具有足够的稳健性。然而，某些参数的影响因细胞系而异，这表明对于每个感兴趣的细胞系都应验证Cedex的性能。
