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Use of the 1-mm micro-probe for metabolic analysis on small volume biological samples.

Serkova NJ, Freund AS, Brown JL, Kominsky DJ - J. Cell. Mol. Med. (2009)

Bottom Line: Clinical application of high-resolution NMR spectroscopy is often limited by extremely low volumes of human specimens.In the present study, the use of the Bruker 1-mm high-resolution TXI micro-probe was evaluated in the elucidation of metabolic profiles for three different clinical applications with limited sample sizes (body fluids, isolated cells and tissue biopsies).In this study, the use of the Bruker 1-mm micro-probe provides a convenient way to measure and quantify endogenous metabolic profiles of samples with a very low volume/weight/cell count.

View Article: PubMed Central - PubMed

Affiliation: Biomedical MRI/MRS Cancer Center Core, University of Colorado Health Sciences Center, Denver, CO 80262, USA.

ABSTRACT
Endogenous metabolites are promising diagnostic end-points in cancer research. Clinical application of high-resolution NMR spectroscopy is often limited by extremely low volumes of human specimens. In the present study, the use of the Bruker 1-mm high-resolution TXI micro-probe was evaluated in the elucidation of metabolic profiles for three different clinical applications with limited sample sizes (body fluids, isolated cells and tissue biopsies). Sample preparation and (1)H-NMR metabolite quantification protocols were optimized for following oncology-oriented applications: (i) to validate the absolute concentrations of citrate and spermine in human expressed prostatic specimens (EPS volumes 5 to 10 microl: prostate cancer application); (ii) to establish the metabolic profile of isolated human lymphocytes (total cell count 4 x 10(6): chronic myelogenous leukaemia application); (iii) to assess the metabolic composition of human head-and-neck cancers from mouse xenografts (biopsy weights 20 to 70 mg: anti-cancer treatment application). In this study, the use of the Bruker 1-mm micro-probe provides a convenient way to measure and quantify endogenous metabolic profiles of samples with a very low volume/weight/cell count.

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1H‐NMR spectrum of 20 mM metabolite solution in D2O supplemented with TMSP (ca. 0.05 wt%) for precise TMSP concentration calculations.
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f1: 1H‐NMR spectrum of 20 mM metabolite solution in D2O supplemented with TMSP (ca. 0.05 wt%) for precise TMSP concentration calculations.

Mentions: All 2D data were processed using XWINNMR 3.5 or TopSpin software. Tissue metabolites were identified based on the results from our chemical shift database and/or referred to the Human Metabolome Database from the University of Alberta (http://www.hmdb.ca/). After performing Fourier transformation (with line broadening LB = 0.2 Hz) on one‐dimensional 1H‐NMR spectra and making phase and baseline corrections, each identified 1H peak was integrated using Bruker 1D WINNMR program. The final TMSP concentration in the capillary was calculated prior to metabolite calculations in the study extracts using a 20 mM metabolite standard solution. TMSP concentrations were calculated based on NMR intensities (from the fully relaxed 1H‐NMR spectra) of each metabolite according to equation (1) (Fig. 1) and the final concentration of TMSP (given as mmol/ml) represented a mean value (Table 1).


Use of the 1-mm micro-probe for metabolic analysis on small volume biological samples.

Serkova NJ, Freund AS, Brown JL, Kominsky DJ - J. Cell. Mol. Med. (2009)

1H‐NMR spectrum of 20 mM metabolite solution in D2O supplemented with TMSP (ca. 0.05 wt%) for precise TMSP concentration calculations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4940789&req=5

f1: 1H‐NMR spectrum of 20 mM metabolite solution in D2O supplemented with TMSP (ca. 0.05 wt%) for precise TMSP concentration calculations.
Mentions: All 2D data were processed using XWINNMR 3.5 or TopSpin software. Tissue metabolites were identified based on the results from our chemical shift database and/or referred to the Human Metabolome Database from the University of Alberta (http://www.hmdb.ca/). After performing Fourier transformation (with line broadening LB = 0.2 Hz) on one‐dimensional 1H‐NMR spectra and making phase and baseline corrections, each identified 1H peak was integrated using Bruker 1D WINNMR program. The final TMSP concentration in the capillary was calculated prior to metabolite calculations in the study extracts using a 20 mM metabolite standard solution. TMSP concentrations were calculated based on NMR intensities (from the fully relaxed 1H‐NMR spectra) of each metabolite according to equation (1) (Fig. 1) and the final concentration of TMSP (given as mmol/ml) represented a mean value (Table 1).

Bottom Line: Clinical application of high-resolution NMR spectroscopy is often limited by extremely low volumes of human specimens.In the present study, the use of the Bruker 1-mm high-resolution TXI micro-probe was evaluated in the elucidation of metabolic profiles for three different clinical applications with limited sample sizes (body fluids, isolated cells and tissue biopsies).In this study, the use of the Bruker 1-mm micro-probe provides a convenient way to measure and quantify endogenous metabolic profiles of samples with a very low volume/weight/cell count.

View Article: PubMed Central - PubMed

Affiliation: Biomedical MRI/MRS Cancer Center Core, University of Colorado Health Sciences Center, Denver, CO 80262, USA.

ABSTRACT
Endogenous metabolites are promising diagnostic end-points in cancer research. Clinical application of high-resolution NMR spectroscopy is often limited by extremely low volumes of human specimens. In the present study, the use of the Bruker 1-mm high-resolution TXI micro-probe was evaluated in the elucidation of metabolic profiles for three different clinical applications with limited sample sizes (body fluids, isolated cells and tissue biopsies). Sample preparation and (1)H-NMR metabolite quantification protocols were optimized for following oncology-oriented applications: (i) to validate the absolute concentrations of citrate and spermine in human expressed prostatic specimens (EPS volumes 5 to 10 microl: prostate cancer application); (ii) to establish the metabolic profile of isolated human lymphocytes (total cell count 4 x 10(6): chronic myelogenous leukaemia application); (iii) to assess the metabolic composition of human head-and-neck cancers from mouse xenografts (biopsy weights 20 to 70 mg: anti-cancer treatment application). In this study, the use of the Bruker 1-mm micro-probe provides a convenient way to measure and quantify endogenous metabolic profiles of samples with a very low volume/weight/cell count.

Show MeSH
Related in: MedlinePlus