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Validation of a Dye Stain Assay for Vaginally Inserted Hydroxyethylcellulose-Filled Microbicide Applicators

Katzen, Lauren L. MPA*; Fernández-Romero, José A. MS*; Sarna, Avina MD MPH, PhD; Murugavel, Kailapuri G. MSc, PhD; Gawarecki, Daniel MA*; Zydowsky, Thomas M. PhD*; Mensch, Barbara S. PhD*

doi: 10.1097/OLQ.0b013e31822e6160
Original Study

Background: The reliability and validity of self-reports of vaginal microbicide use are questionable given the explicit understanding that participants are expected to comply with study protocols. Our objective was to optimize the use of Population Council's previously validated dye stain assay (DSA) and related procedures, and to establish predictive values for the DSAs ability to identify vaginally inserted single-use, low-density polyethylene microbicide applicators filled with hydroxyethylcellulose gel.

Methods: Applicators, inserted by 252 female sex workers enrolled in a microbicide feasibility study in Southern India, served as positive controls for optimization and validation experiments. Before validation, optimal dye concentration and staining time were ascertained. Three validation experiments were conducted to determine sensitivity, specificity, and negative and positive predictive values.

Results: The dye concentration of 0.05% (wt/vol) FD&C Blue No. 1 Granular Food Dye (Prime Ingredients, Inc, Saddlebrook, NJ) and staining time of 5 seconds were determined to be optimal and were used for the 3 validation experiments. There were a total of 1848 possible applicator readings across validation experiments; 1703 (92.2%) applicator readings were correct. On average, the DSA performed with 90.6% sensitivity, 93.9% specificity, and had a negative predictive value of 93.8% and a positive predictive value of 91.0%. No statistically significant differences between experiments were noted.

Conclusions: The DSA was optimized and successfully validated for use with single use, low-density polyethylene applicators filled with hydroxyethylcellulose gel. We recommend including the DSA in future microbicide trials involving vaginal gels so as to identify participants who have low adherence to dosing regimens. In doing so, we can develop strategies to improve adherence as well as investigate the association between product use and efficacy.

The optimization and validation of the Population Council's dye stain assay to identify vaginal insertion of low-density polyethylene microbicide applicators filled with hydroxyethylcellulose gel.

From the *Population Council, New York, NY; †Population Council, New Delhi, India; and ‡YRG CARE (Y.R. Gaitonde Center for AIDS Research and Education), Chennai, India

The authors would like to thank Meredith Paddock and Barbara Miller for their indispensable assistance in the preparation of this manuscript, and Sharon Abbott, Barbara Friedland, Sarah Littlefield, Deborah Tolenaar, and Waimar Tun for their insightful comments, guidance, and support. They would also like to thank C. Chellamani, C. Shalini, C. Sivanesan, Madhan, and V. Vijaya for their dedicated work on the India study.

This manuscript is made possible by the generous support of the American and Indian people through the United States Agency for International Development (USAID), Bureau for Global Health, Office of Population and Reproductive Health, under the terms of Award No. GPO-A-00-04-00019, the Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), under the terms of Award No. R21 HD060270, and the Indian Council of Medical Research (ICMR), under the terms of Award No. Indo-US/54/2007-ECD II.

The contents of this manuscript are the sole responsibility of the Population Council and do not necessarily reflect the views of USAID, NIH, or the United States Government.

Correspondence: Lauren L. Katzen, MPA, Center for Biomedical Research, 1230 York Avenue, New York, NY 10065. E-mail: lkatzen@popcouncil.org.

Received for publication March 24, 2011, and accepted July 18, 2011.

© Copyright 2011 American Sexually Transmitted Diseases Association