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Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses
Version 1
: Received: 22 May 2024 / Approved: 22 May 2024 / Online: 23 May 2024 (08:19:18 CEST)
How to cite:
Kaneko, S.; Okada, Y. Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses. Preprints2024, 2024051442. https://doi.org/10.20944/preprints202405.1442.v1
Kaneko, S.; Okada, Y. Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses. Preprints 2024, 2024051442. https://doi.org/10.20944/preprints202405.1442.v1
Kaneko, S.; Okada, Y. Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses. Preprints2024, 2024051442. https://doi.org/10.20944/preprints202405.1442.v1
APA Style
Kaneko, S., & Okada, Y. (2024). Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses. Preprints. https://doi.org/10.20944/preprints202405.1442.v1
Chicago/Turabian Style
Kaneko, S. and Yuki Okada. 2024 "Revalidation of DNA Fragmentation Analyses for Human Sperm—Measurement Principles, Comparative Standards, Calibration Curve, Required Sensitivity, and Eligibility Criteria for Test Sperm—A Step-by-Step Approach to Cohort Studies with Multivariable Analyses" Preprints. https://doi.org/10.20944/preprints202405.1442.v1
Abstract
(1) Background: Double-strand breaks (DSBs) in a single nucleus are usually measured using the sperm chromatin structure assay (SCSA), sperm chromatin dispersion (SCD) test, and comet assay (CA). We developed mono-dimensional single-cell pulsed-field gel electrophoresis (1D-SCPFGE) and angle-modulated two-dimensional single-cell pulsed-field gel electrophoresis (2D-SCPFGE) to observe DNA fragmentation in separated motile sperm. (2) Methods: We set up comparative standards, calibration curves, the required sensitivity, and eligibility criteria for test sperm to validate their measurement principles. (3) Results: We revealed that the conventional methods overlooked the interference of nucleoproteins in their measurements. In-gel proteolysis improves the measurement accuracies of 1D- and 2D-SCPFGE. We recommend using naked DNA for comparative standards and test specimens. Moreover, we have observed several dysfunctions that may induce DNA damage in the separated motile sperm. Overall, our discussion highlights the need to revisit the conventional univariable analyses based on the SCSA, SCD test, and CA.
(4) Conclusions: Human infertility is a complex syndrome, and the aim of quality control in intracytoplasmic sperm injection is to identify the underlying dysfunctions remaining in the separated motile sperm that render them ineligible for injection. Well-designed, multivariable analyses with special consideration to confounding factors are necessary in future cohort studies.
Keywords
DNA fragmentation; single-cell pulsed-field gel electrophoresis; human sperm; sperm chromatin structure assay; sperm chromatin dispersion test; comet assay; quantitative analysis
Subject
Biology and Life Sciences, Biology and Biotechnology
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
