Effects of Repeated Freeze-Thaw Cycles on Concentrations of Cholesterol, Micronutrients, and Hormones in Human Plasma and Serum

Abstract

The effects of repeated freeze-thaw cycles on concentrations of various analytes in plasma or serum were of little interest until the growth of plasma and serum banks during the latter part of the 20th century. By 1996, the number of such banks used primarily for cancer research had grown to 115 (1). Many also exist for other purposes, such as the WHO Serum Reference Banks (2) and banks associated with cardiovascular studies such as the Multiple Risk Factor Intervention Trial (MRFIT) and the Atherosclerosis Risk in Communities (ARIC) Study (3)(4). Although the need for repeated freezing and thawing of samples can be minimized by storing banked specimens in several small containers (5), it often is necessary to use plasma or serum that has already undergone one or more freeze-thaw cycles. When this occurs, reviewers of research protocols or manuscripts may question the validity of data obtained from these specimens. The scientific literature provides few answers to such questions. Medline contains no appropriate key word or phrase for searching; comments on the effects of freeze-thaw cycles often are limited to a few sentences in publications that focus on stability during long-term storage or on assay methodology. The present study was designed to add information to this sparse literature. In 1991, 10 healthy adult volunteers, 5 men and 5 women, each donated 120 mL of blood. The purpose of the study was explained to each person along with the risks and lack of individual benefit. Approval of the study was granted by the Committee on Human Volunteers of the Johns Hopkins School of Hygiene and Public Health before study initiation. Blood was collected into three 20-mL Vacutainer Tubes containing 286 USP units of sodium heparin (cat. no. 6406; Becton Dickinson) to obtain plasma, and into four 15-mL plain Vacutainer Tubes …