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Effect of multiple freeze-thaw cycles on selected biochemical serum components.

Author(s): Gislefoss RE, Lauritzen M, Langseth H, Mørkrid L

Publication: Clin Chem Lab Med, 2017, Vol. 55(7), Page 967-973

PubMed ID: 27987362 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study investigated the effects of up to ten freeze-thaw cycles (1–10) on 15 biochemical analytes in serum. Peripheral blood was collected from 40 fasting volunteers aged 30–60 years (health status not stated) in Vacutainer tubes and left to coagulate for 1 h prior to centrifugation for 10 min at 1300 × g. Serum was aliquoted and then frozen at -25°C. Once a week, specimens were thawed at room temperature, mixed for 10 min on a blood sample roller, and then re-frozen by returning to -25˚C freezer. Specimens obtained after 1, 2, 3, 4, 5, or 10 freeze-thaw cycles were analyzed in the same batch after three months of storage. Albumin, aspartate-aminotransferase (ASAT), cholesterol, glucose, immunoglobulin G, triglycerides, urea, bilirubin, potassium, sodium, levels were measured on an automated analyzer (enzymatic, colorimetric, immunoturbidimetric, or ion selective electrode method); testosterone, thyroid stimulating hormone (TSH), and vitamin B12 were assayed by an electrochemiluminescence immunoassay; creatinine levels were determined using an enzymatic assay; and CRP levels determined by an immunoturbidimetric assay. Twenty specimens thawed after 24 h frozen storage were compared with fresh specimens. Analyte levels for the remaining freeze-thaw groups were compared to the first thaw group results.

   Summary of Findings:

   Levels of only five serum analytes were statistically significant from those in fresh specimens after one freeze-thaw cycle (bilirubin P<0.001, cholesterol P=0.014, IgG P= 0.037, TG P<0.001, and urea P=0.001). While albumin, ASAT, bilirubin, potassium, sodium, testosterone, and TSH showed within-specimen statistically significant differences due to repeated freeze-thaw cycles, only levels of bilirubin, sodium, and TSH were significantly affected in the pairwise comparisons to first freeze-thaw cycle levels (bilirubin P<0.001 ≥2 freeze-thaw cycles, sodium P=0.01 after 4 freeze-thaw cycles, and TSH P<0.001 after ≥5 freeze-thaw cycles and P=0.019 after 10 freeze-thaw cycles). However, differences observed for albumin after 5 freeze-thaw cycles; ASAT and testosterone after 10 cycles; bilirubin after ≥3, cycles; and sodium after ≥2 freeze-thaw cycles were considered clinically significant.

   Biospecimens

   • Bodily Fluid - Serum

   Preservative Types

   • Frozen
   • None (Fresh)

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   Electrolyte/Metal	Ion selective electrode
   Protein	Immunoturbidometric assay
   Protein	Enzyme assay
   Protein	Chemiluminescence immunoassay
   Protein	Clinical chemistry/auto analyzer
   Steroid	Clinical chemistry/auto analyzer
   Small molecule	Clinical chemistry/auto analyzer
   Glycoprotein	Clinical chemistry/auto analyzer

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Freeze/thaw cycling	1 cycle 2 cycles 3 cycles 4 cycles 5 cycles 10 cycles
   Biospecimen Preservation	Type of fixation/preservation	Frozen None (fresh)

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