Analysis of Subforms of Free Prostate-Specific Antigen in Serum by Two-Dimensional Gel Electrophoresis: Potential to Improve Diagnosis of Prostate Cancer

Jung, Klaus; Reiche, Janett; Boehme, Axel; Stephan, Carsten; Loening, S.A.; Schnorr, Dietmar; Hoesel, Wolfgang; Sinha, Pranav

doi:10.1373/clinchem.2004.040469

Cited by 28 publications

(33 citation statements)

References 47 publications

(52 reference statements)

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“…The two PSA+ samples each show three major PSA isoforms falling within the pI 6.4-7.5 range, in good agreement with comparatively laborious slab-gel IEF studies (6,8). Patient-specific differences in PSA isoform representation and pI are clearly apparent, recapitulating the potential utility of isoform ratio measurements in clinical diagnostics and personalized medicine (6,7).…”

Section: Microfluidic Lavagel Analysis Of Psa Isoforms In Metastatic supporting

confidence: 63%

“…Improvement of the absolute lower limit of detection should be feasible through, for example, incorporation of amplified readout approaches to yield a similar sensitivity to conventional benchtop 2D electrophoresis with Western blotting (ca. 0.1 ng mL −1 ) (6,19). Such approaches should also increase the dynamic operating range of the present assay.…”

Section: Resultsmentioning

confidence: 94%

“…In one important example, the mediocre ability of total and free PSA assays to distinguish between malignant and benign prostatic pathology has spurred study of free PSA isoforms (6,7). Proteomic studies suggest a promising link between prostate cancer incidence and differential isoform expression in healthy and cancer patient sera (6,8,9).…”

mentioning

confidence: 99%

“…Proteomic studies suggest a promising link between prostate cancer incidence and differential isoform expression in healthy and cancer patient sera (6,8,9). Although promising, rigorous validation studies are needed to translate the potential of protein isoforms to the clinic.…”

mentioning

confidence: 99%

“…Recent ELISA-based formats offer notable gains in analytical sensitivity (10,11). Nevertheless, mounting evidence suggests that protein isoform "fingerprinting" could advance diagnostic performance (6,9,12). Unfortunately, ELISA is severely limited for isoform discrimination because antibodies specific to protein isoforms often do not exist (12).…”

mentioning

confidence: 99%

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Microfluidic integration for automated targeted proteomic assays

Hughes

Lin

Peehl

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

103

167

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A dearth of protein isoform-based clinical diagnostics currently hinders advances in personalized medicine. A well-organized protein biomarker validation process that includes facile measurement of protein isoforms would accelerate development of effective protein-based diagnostics. Toward scalable protein isoform analysis, we introduce a microfluidic "single-channel, multistage" immunoblotting strategy. The multistep assay performs all immunoblotting steps: separation, immobilization of resolved proteins, antibody probing of immobilized proteins, and all interim wash steps. Programmable, low-dispersion electrophoretic transport obviates the need for pumps and valves. A three-dimensional bulk photoreactive hydrogel eliminates manual blotting. In addition to simplified operation and interfacing, directed electrophoretic transport through our 3D nanoporous reactive hydrogel yields superior performance over the state-of-the-art in enhanced capture efficiency (on par with membrane electroblotting) and sparing consumption of reagents (ca. 1 ng antibody), as supported by empirical and by scaling analyses. We apply our fully integrated microfluidic assay to protein measurements of endogenous prostate specific antigen isoforms in (i) minimally processed human prostate cancer cell lysate (1.1 pg limit of detection) and (ii) crude sera from metastatic prostate cancer patients. The single-instrument functionality establishes a scalable microfluidic framework for high-throughput targeted proteomics, as is relevant to personalized medicine through robust protein biomarker verification, systematic characterization of new antibody probes for functional proteomics, and, more broadly, to characterization of human biospecimen repositories.isoelectric focusing | nanoporous reactive polymers | lab-on-a-chip | Western blotting | antibody selection

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Section: Microfluidic Lavagel Analysis Of Psa Isoforms In Metastatic supporting

confidence: 63%

Section: Resultsmentioning

confidence: 94%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

Microfluidic integration for automated targeted proteomic assays

Hughes

Lin

Peehl

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

103

167

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Comparative analysis of prostate‐specific antigen by two‐dimensional gel electrophoresis and capillary electrophoresis

et al. 2016

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Serum levels of Prostate-Specific Antigen (PSA) are not fully specific for prostate cancer (PCa) diagnosis and several efforts are focused on searching to improve PCa markers through the study of PSA subforms that could be cancer associated. We have previously reported by 2DE a decrease in the sialic acid content of PSA from PCa compared to benign prostatic hyperplasia patients based on the different proportion of the PSA spots. However, faster and more quantitative techniques, easier to automate than 2DE, are desirable. In this study, we examined the potential of CE for resolving PSA subforms in different samples and compared the results with those obtained by 2DE. We first fractionated by OFFGEL the subforms of PSA from seminal plasma according to their pIs and analyzed each separated fraction by 2DE and CE. We also analyzed PSA and high pI PSA, both from seminal plasma, and PSA from urine of a PCa patient. These samples with different PSA spots proportions by 2DE, due to different posttranslational modifications, also presented different CE profiles. This study shows that CE is a useful and complementary technique to 2DE for analyzing samples with different PSA subforms, which is of high clinical interest.

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Human body fluid proteome analysis

2006

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The focus of this article is to review the recent advances in proteome analysis of human body fluids, including plasma/serum, urine, cerebrospinal fluid, saliva, bronchoalveolar lavage fluid, synovial fluid, nipple aspirate fluid, tear fluid, and amniotic fluid, as well as its applications to human disease biomarker discovery. We aim to summarize the proteomics technologies currently used for global identification and quantification of body fluid proteins, and elaborate the putative biomarkers discovered for a variety of human diseases through human body fluid proteome (HBFP) analysis. Some critical concerns and perspectives in this emerging field are also discussed. With the advances made in proteomics technologies, the impact of HBFP analysis in the search for clinically relevant disease biomarkers would be realized in the future.

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Analysis of Subforms of Free Prostate-Specific Antigen in Serum by Two-Dimensional Gel Electrophoresis: Potential to Improve Diagnosis of Prostate Cancer

Cited by 28 publications

References 47 publications

Microfluidic integration for automated targeted proteomic assays

Microfluidic integration for automated targeted proteomic assays

Comparative analysis of prostate‐specific antigen by two‐dimensional gel electrophoresis and capillary electrophoresis

Human body fluid proteome analysis

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