期刊
CELLS
卷 4, 期 1, 页码 21-39出版社
MDPI
DOI: 10.3390/cells4010021
关键词
ELISPOT; Smart Count (TM); Log Normal distribution; harmonization
类别
资金
- Novo Nordisk Fonden [NNF14OC0012533, NNF12OC0002037] Funding Source: researchfish
The primary goal of immune monitoring with ELISPOT is to measure the number of T cells, specific for any antigen, accurately and reproducibly between different laboratories. In ELISPOT assays, antigen-specific T cells secrete cytokines, forming spots of different sizes on a membrane with variable background intensities. Due to the subjective nature of judging maximal and minimal spot sizes, different investigators come up with different numbers. This study aims to determine whether statistics-based, automated size-gating can harmonize the number of spot counts calculated between different laboratories. We plated PBMC at four different concentrations, 24 replicates each, in an IFN-gamma ELISPOT assay with HCMV pp65 antigen. The ELISPOT plate, and an image file of the plate was counted in nine different laboratories using ImmunoSpot (R) Analyzers by (A) Basic Count T relying on subjective counting parameters set by the respective investigators and (B) SmartCount (TM), an automated counting protocol by the ImmunoSpot (R) Software that uses statistics-based spot size auto-gating with spot intensity auto-thresholding. The average coefficient of variation (CV) for the mean values between independent laboratories was 26.7% when counting with Basic Count (TM), and 6.7% when counting with SmartCount (TM). Our data indicates that SmartCount (TM) allows harmonization of counting ELISPOT results between different laboratories and investigators.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据