Quality assurance considerations in cryptosporidium antibody tests.

Abstract

Priest et al. (12) reported a comparison of serological techniques to detect responses to two Cryptosporidium antigens. They compared a low-cost minigel format blot assay that we developed with a Centers for Disease Control and Prevention (CDC)-developed enzyme-linked immunosorbent assay (ELISA) and a large-format gel. The minigel analysis was performed in a British Columbia laboratory. The minigel and large-format gel detected similar positive and negative responses to the 15- and 17-kDa antigens; however, the CDC ELISA had 23% false-positive responses and 12% false-negative responses to the 15- to 17-kDa antigen. For the 27-kDa antigen, the large-format gel detected more positive responses than did either the minigel or ELISA. In 1996, we trained the British Columbia staff in Albuquerque to conduct our minigel assay and provided copies of our quality assurance (QA) procedures. These procedures require computer imaging of the blots and rejecting blots with a weak positive control (PC) or when the coefficient of variation (CV) for duplicate PCs is greater than 35% (3). When contacted, Dr. Priest stated that the blots used in their publication were not computer imaged and that QA procedures were informally conducted by manual inspection. Although we requested computer images of these blots for additional QA evaluation, they were not made available to us. To reduce costs, the minigel uses only a small fraction of the antigen needed for either the ELISA or large-format blot. As a result, considerable care must be taken to minimize sources of variance and to insure that the antigen is uniformly applied across the blot. In addition, since we are interested in both detecting a response and measuring the intensity of that response, it is critical that blot performance be continuously monitored. Since responses tend to fade over time, the imaging must be done shortly after the blots are completed to avoid underestimating the CV. Because the cost of the minigel assay is dramatically lower than the cost of the large-format gel assay and, we believe, lower than the cost of the ELISA, we compared performance of the minigel to that of the large-format gel. The large-format gel assay was used as the standard of comparison by Priest et al. We compared detection of a response to each antigen for 42 samples from two prior studies (3, 4). Concordance in detection was 79% for the 15- to 17-kDa antigen and 86% for the 27-kDa antigen. The minigel detected six responses to the 15to 17-kDa antigen and three to the 27-kDa antigen that the large gel missed. The large gel detected three responses to the 15- to 17-kDa antigen and three to the 27-kDa antigen that the minigel missed. These data indicate that the minigel assay, with appropriate quality control procedures, can generate findings comparable to those for the large-format gel assay at a small fraction of the cost.