BACKGROUND: Beyond determining the percentage of damaged sperm, current methods of DNA assessment are of limited clinical utility as they render the sample unusable. We evaluated Raman microspectroscopy, a laser-based non-invasive technique that provides detailed chemical 'fingerprints' of cells and which potentially could be used for nuclear DNA-based sperm selection. METHODS: Eight healthy donors provided ejaculates. After system optimization, a minimum of 200 air-dried sperm/sample/donor, prior to/and after UVB irradiation, were assessed by two observers. Spectra were analysed by Principal Component, Spectral Angle and Wavelet Analyses. RESULTS: Spectra provided a chemical map delineating each sperm head region. Principal Component Analysis showed clear separation between spectra from UV-irradiated and untreated samples whilst averaged data identified two regions of interest (1040 and 1400 cm(-1)). Local spectral analysis around the DNA PO(4) backbone peak (1042 cm(-1)), showed that changes in this region were indicative of DNA damage. Wavelet decomposition confirmed both the 1042 cm(-1) shift and a second UVB susceptible region (1400-1600 cm(-1)) corresponding to protein-DNA interactions. No difference was found between observer measurements. CONCLUSIONS: Raman microspectroscopy can provide accurate and reproducible assessment of sperm DNA structure and the sites and location of damage.
BACKGROUND: Beyond determining the percentage of damaged sperm, current methods of DNA assessment are of limited clinical utility as they render the sample unusable. We evaluated Raman microspectroscopy, a laser-based non-invasive technique that provides detailed chemical 'fingerprints' of cells and which potentially could be used for nuclear DNA-based sperm selection. METHODS: Eight healthy donors provided ejaculates. After system optimization, a minimum of 200 air-dried sperm/sample/donor, prior to/and after UVB irradiation, were assessed by two observers. Spectra were analysed by Principal Component, Spectral Angle and Wavelet Analyses. RESULTS: Spectra provided a chemical map delineating each sperm head region. Principal Component Analysis showed clear separation between spectra from UV-irradiated and untreated samples whilst averaged data identified two regions of interest (1040 and 1400 cm(-1)). Local spectral analysis around the DNA PO(4) backbone peak (1042 cm(-1)), showed that changes in this region were indicative of DNA damage. Wavelet decomposition confirmed both the 1042 cm(-1) shift and a second UVB susceptible region (1400-1600 cm(-1)) corresponding to protein-DNA interactions. No difference was found between observer measurements. CONCLUSIONS: Raman microspectroscopy can provide accurate and reproducible assessment of sperm DNA structure and the sites and location of damage.
Authors: Fa-Ke Lu; Srinjan Basu; Vivien Igras; Mai P Hoang; Minbiao Ji; Dan Fu; Gary R Holtom; Victor A Neel; Christian W Freudiger; David E Fisher; X Sunney Xie Journal: Proc Natl Acad Sci U S A Date: 2015-08-31 Impact factor: 11.205
Authors: Viviane Ribas Pereira; Danillo Roberto Pereira; Kátia Cristina de Melo Tavares Vieira; Vitor Pereira Ribas; Carlos José Leopoldo Constantino; Patrícia Alexandra Antunes; Ana Paula Alves Favareto Journal: Environ Sci Pollut Res Int Date: 2019-11-07 Impact factor: 4.223