Dataset Information

Quantitation of peptides from non-invasive skin tapings using isotope dilution and tandem mass spectrometry.

ABSTRACT: Previous work from our laboratories utilized a novel skin taping method and mass spectrometry-based proteomics to discover clinical biomarkers of skin conditions; these included atopic dermatitis, Staphylococcus aureus colonization, and eczema herpeticum. While suitable for discovery purposes, semi-quantitative proteomics is generally time-consuming and expensive. Furthermore, depending on the method used, discovery-based proteomics can result in high variation and inadequate sensitivity to detect low abundant peptides. Therefore, we strove to develop a rapid, sensitive, and reproducible method to quantitate disease-related proteins from skin tapings. We utilized isotopically-labeled peptides and tandem mass spectrometry to obtain absolute quantitation values on 14 peptides from 7 proteins; these proteins had shown previous importance in skin disease. The method demonstrated good reproducibility, dynamic range, and linearity (R2?>?0.993) when n?=?3 standards were analyzed across 0.05-2.5?pmol. The method was used to determine if differences exist between skin proteins in a small group of atopic versus non-atopic individuals (n?=?12). While only minimal differences were found, peptides were detected in all samples and exhibited good correlation between peptides for 5 of the 7 proteins (R2?=?0.71-0.98). This method can be applied to larger cohorts to further establish the relationships of these proteins to skin disease.

SUBMITTER: Reisdorph N

PROVIDER: S-EPMC6093185 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Quantitation of peptides from non-invasive skin tapings using isotope dilution and tandem mass spectrometry.

Reisdorph Nichole N Armstrong Michael M Powell Roger R Quinn Kevin K Legg Kevin K Leung Donald D Reisdorph Rick R

Journal of chromatography. B, Analytical technologies in the biomedical and life sciences 20180321

Previous work from our laboratories utilized a novel skin taping method and mass spectrometry-based proteomics to discover clinical biomarkers of skin conditions; these included atopic dermatitis, Staphylococcus aureus colonization, and eczema herpeticum. While suitable for discovery purposes, semi-quantitative proteomics is generally time-consuming and expensive. Furthermore, depending on the method used, discovery-based proteomics can result in high variation and inadequate sensitivity to dete ...[more]

PMID: 29601982

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Project description:Castration resistant prostate cancer (CRPC), the fatal form of prostate cancer, remains androgen dependent despite castrate levels of circulating testosterone (T) and 5α-dihydrotestosterone (DHT). To investigate mechanisms by which the tumor can synthesize its own androgens and develop resistance to abiraterone acetate and enzalutamide, methods to measure a complete androgen profile are imperative. Here, we report the development and validation of a stable isotope dilution liquid chromatography electrospray ionization tandem mass spectrometric (SID-LC-ESI-MS/MS) method to quantify nine human hydroxy-androgens as picolinates, simultaneously with requisite specificity and sensitivity. In the established method, the fragmentation patterns of all nine hydroxy-androgen picolinates were identified, and [13C3]-5α-androstane-3α, 17β-diol and [13C3]-5α-androstane-3β, 17β-diol used as internal standards were synthesized enzymatically. Intra-day and inter-day precision and accuracy corresponds to the U.S. Food and Drug Administration Criteria for Bioanalytical Method Validation. The lower limit of quantitation (LLOQ) of nine hydroxy-androgens is 1.0pg to 2.5pg on column. Diols which have been infrequently measured: 5-androstene-3β, 17β-diol and 5α-androstane-3α, 17β-diol can be determined in serum at values as low as 1.0pg on column. The method also permits the quantitation of conjugated hydroxy-androgens following enzymatic digestion. While direct detection of steroid conjugates by electrospray-ionization tandem mass spectrometry has advantages the detection of unconjugated and conjugated steroids would require separate methods for each set of analytes. Our method was applied to pooled serum from male and female donors to provide reference values for both unconjugated and conjugated hydroxy-androgens. This method will allow us to interrogate the involvement of the conversion of 5-androstene-3β, 17β-diol to T, the backdoor pathway involving the conversion of 5α-androstane-3α, 17β-diol to DHT and the inactivation of DHT to 5α-androstane-3β, 17β-diol in advanced prostate cancer.

Dataset Information

Quantitation of peptides from non-invasive skin tapings using isotope dilution and tandem mass spectrometry.

Publications

Quantitation of peptides from non-invasive skin tapings using isotope dilution and tandem mass spectrometry.

Similar Datasets

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