Victoria Sánchez, B. Sc. , Klaus Redmann, Ph. D. , Joachim Wistuba, Ph

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Presentation transcript:

Oxidative DNA damage in human sperm can be detected by Raman microspectroscopy Victoria Sánchez, B.Sc., Klaus Redmann, Ph.D., Joachim Wistuba, Ph.D., Frank Wübbeling, Ph.D., Martin Burger, Ph.D., Harriette Oldenhof, Ph.D., Willem F. Wolkers, Ph.D., Sabine Kliesch, M.D., Stefan Schlatt, Ph.D., Con Mallidis, Ph.D. Fertility and Sterility Volume 98, Issue 5, Pages 1124-1129.e3 (November 2012) DOI: 10.1016/j.fertnstert.2012.07.1059 Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Figure 1 Flow-cytometric assessment of oxidative sperm nDNA damage showing the four different levels of fragmentation induced by Fenton's reaction. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Figure 2 (A) Raman spectra of an untreated spermatozoon with 27% DFI (black) and of another after induced oxidative nDNA damage resulting in 92% DFI (red). (B) FTIR spectra of pellets of the same samples (i.e., same DFI). Arrows in both cases indicate the peaks where changes associated with nDNA damage were found. The intensities of these bands and their relative ratios are detailed in the embedded tables. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Figure 3 Overlay of mean Raman (A) and FTIR (B) spectra in the region corresponding to the DNA PO4 backbone indicating different levels of oxidative damage. Spectra have been separated for clarity. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Figure 4 Correlation of band intensity ratios with DFI values of the mean spectra from both Raman (A) and FTIR (B) showed significant degrees of linearity with the results of the flow-cytometric assessment. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 1 PCA of Raman spectra over the range 600–1,800 cm−1. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 2 Band intensity ratios (1,050/1,095 cm−1) of Raman spectra from 200 sperm per sample indicating presence of nDNA fragmentation. Computation of a cutoff value of 0.63 (dotted line) allowed for the later determination of the percentage of sperm with damage present in the sample. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 3 (A) Correlation of DFI and the percent of sperm with peak intensity ratio 1,050/1,095 cm−1 higher than the determined cutoff value (0.63). (B) Correlation of secondary peak intensity ratios (1,446/785 cm−1) and DFI values from Raman mean spectra. Fertility and Sterility 2012 98, 1124-1129.e3DOI: (10.1016/j.fertnstert.2012.07.1059) Copyright © 2012 American Society for Reproductive Medicine Terms and Conditions