研究目的
Defining the pathogenesis of renal osteodystrophy (ROD) and its treatment efficacy, and identifying differences in bone composition between healthy and ROD bone tissues using confocal Raman microscopy and statistical analysis.
研究成果
Confocal Raman microscopy combined with statistical analysis effectively distinguishes between healthy and ROD bone tissues by identifying significant differences in compositional ratios (calcium content, mineral-to-matrix, carbonate-to-matrix) and observing elevated phenylalanine levels in ROD samples for the first time. The method requires a relatively small number of spectra for accurate classification, supporting the potential for future in vivo Raman sensor development for non-invasive bone quality assessment.
研究不足
The study is based on in vitro measurements of embedded bone samples, which may not fully replicate in vivo conditions. The method requires specialized equipment and expertise, and the sample size is relatively small (10 specimens). Additionally, the collagen matrix analysis in the amide I region is less precise compared to FTIR measurements, and a larger number of spectra may be needed for certain analyses.
1：Experimental Design and Method Selection:
The study employed confocal Raman microscopy to analyze bone tissue composition, focusing on parametric ratios such as calcium content, mineral-to-matrix, and carbonate-to-matrix. Statistical analysis was used to identify significant differences between healthy and ROD samples.
2：Sample Selection and Data Sources:
10 specimens (5 ROD and 5 normal adult human iliac crest bone biopsies) were selected from the Mayo Clinic database, embedded in PMMA, and sectioned into 5-micron thick slices.
3：List of Experimental Equipment and Materials:
A Leica RM 2265 microtome for sectioning, an alpha 300R WITec confocal Raman system with a 532 nm Nd:YAG laser, a 20X Olympus objective lens, and a Marconi 40-11 CCD detector. Software included WITec Control 1.60 and WITec Project Plus for data acquisition and processing, and MATLAB? r2016a for statistical analysis.
4：60 and WITec Project Plus for data acquisition and processing, and MATLAB? r2016a for statistical analysis.
Experimental Procedures and Operational Workflow:
4. Experimental Procedures and Operational Workflow: Raman mapping images were acquired with 150x150 spectra arrays per sample, using an integration time of 50 ms per spectrum and 80 μm x 80 μm scan sizes. Background subtraction, normalization, and outlier isolation were performed before calculating biomarker ratios.
5：Data Analysis Methods:
Statistical analysis included Welch's t-test for comparing average ratios, post-hoc power analysis to determine the number of spectra needed for distinction, and visualization of Raman images for direct assessment of compositional differences.
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