Prostate-specific antigen mRNA and protein levels in laser microdissected cells of human prostate measured by real-time reverse transcriptase–quantitative polymerase chain reaction and immuno–quantitative polymerase chain reaction

Laser-assisted microdissection has mainly been used in cancer studies to excise pure cell populations from heterogeneous tissues. Cancer and normal cells selected by laser-assisted microdissection have frequently been used for mRNA expression studies usually by reverse transcriptase–quantitative pol...

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Bibliographic Details
Published in:Human pathology Vol. 39; no. 10; pp. 1474 - 1482
Main Authors: Pinzani, Pamela, Lind, Kristina, Malentacchi, Francesca, Nesi, Gabriella, Salvianti, Francesca, Villari, Donata, Kubista, Mikael, Pazzagli, Mario, Orlando, Claudio
Format: Journal Article
Language:English
Published: New York, NY Elsevier Inc 01.10.2008
Elsevier
Elsevier Limited
Subjects:
Acids
Adenocarcinoma - genetics
Adenocarcinoma - metabolism
Adenocarcinoma - pathology
Antibodies, Monoclonal - immunology
Biological and medical sciences
Cell Line, Tumor
Enzyme-Linked Immunosorbent Assay
Gene expression
Gene Expression Regulation, Neoplastic
Humans
Hypotheses
Immuno-qPCR
Investigative techniques, diagnostic techniques (general aspects)
Laser microdissection
Lasers
Male
Medical sciences
Methods
Microdissection
Pathology
Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques
Polymerase chain reaction
Polymerase Chain Reaction - methods
Prostate - metabolism
Prostate - pathology
Prostate cancer
Prostate specific antigen
Prostate-Specific Antigen - genetics
Prostate-Specific Antigen - metabolism
Prostatic Neoplasms - genetics
Prostatic Neoplasms - metabolism
Prostatic Neoplasms - pathology
Real time quantitative RT-PCR
Reverse Transcriptase Polymerase Chain Reaction
RNA, Messenger - metabolism
RNA, Neoplasm - analysis
Studies
Immuno-qPCR
Prostate specific antigen
Real time quantitative RT-PCR
Laser microdissection
Human
RNA-directed DNA polymerase
Microdissection
Enzyme
Transferases
Tumoral marker
Real time
Protein
Polymerase chain reaction
Real time quantitative
Nucleotidyltransferases
Anatomic pathology
Messenger RNA
RT-PCR
Laser
Urogenital system
Cell
Prostate
Reverse transcription polymerase chain reaction
Quantitative analysis
ISSN:0046-8177, 1532-8392, 1532-8392
Online Access:Get full text
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Summary:Laser-assisted microdissection has mainly been used in cancer studies to excise pure cell populations from heterogeneous tissues. Cancer and normal cells selected by laser-assisted microdissection have frequently been used for mRNA expression studies usually by reverse transcriptase–quantitative polymerase chain reaction (qPCR). Recently, real time immuno-qPCR was developed as a new tool for highly sensitive measurements of proteins. Using reverse transcriptase–qPCR and immuno-qPCR, we measured the amounts of prostate-specific antigen mRNA and its corresponding protein in homogeneous and comparable cell populations, collected from normal and cancer prostates by laser-assisted microdissection. With these techniques, prostate-specific antigen mRNA and protein were quantified over a wide range of concentrations with a sensitivity sufficient to analyze single prostate cells (LNCaP). We did not find significant differences in prostate-specific antigen protein and mRNA between normal and cancer cells. The expression of prostate-specific antigen protein and mRNA was highly correlated in both normal and pathological cells. In microdissected peritubular stromal areas of prostate cancers, the concentration of prostate-specific antigen protein was about 100 times higher than in normal prostate, indicating an increased transit of secreted prostate-specific antigen. In the same samples, prostate-specific antigen mRNA was not detectable. Our data demonstrate, for the first time, the feasibility of simultaneous application of reverse transcriptase–qPCR and immuno-qPCR in studies of homogeneous cell populations, collected by laser-assisted microdissection. The approach is expected to become a very powerful tool for expression studies in human cancers at both mRNA and protein levels.
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ISSN:0046-8177
1532-8392
1532-8392
DOI:10.1016/j.humpath.2008.02.012