Alexandra A. J. Migchielsen

Leiden University

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Edward J. Goetzl

University of California, San Francisco

Stephan Toutain

Duke University

Hein te Riele

The Netherlands Cancer Institute

Jeffrey A. Golden

Saint Ambrose University

Dinko Valerio

Leiden University Medical Center

Anton Berns

The Netherlands Cancer Institute

Ferry Ossendorp

Leiden University Medical Center

Marian A. van Roon

Amsterdam Neuroscience

Marco L. Breuer

The Netherlands Cancer Institute

Christoph W. Turck

Chinese Academy of Sciences

Cooperative Institutions

Oncode Institute

Leiden University Medical Center

Leiden University

The Netherlands Cancer Institute

Academic Medical Center

University of Amsterdam

University of California, San Francisco

University of California San Francisco Medical Center

Brigham and Women's Hospital

Harvard University

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Adenosine–deaminase–deficient mice die perinatally and exhibit liver–cell degeneration, atelectasis and small intestinal cell death

Nature Genetics (1995)

Alexandra A. J. Migchielsen Marco L. Breuer Marian A. van Roon Hein te Riele C. Zurcher

Deoxyadenosine

Adenosine deaminase deficiency

Transmethylation

Adenosine Deaminase Inhibitor

Deoxycoformycin

5'-nucleotidase

10.1038/ng0795-279

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Citations (117)

Relative quantification of collagen mrna in fibroblasts by a radioactive polymerase chain reaction technique

Journal of Clinical Laboratory Analysis (1991)

Edward J. Goetzl Christoph W. Turck Jeffrey A. Golden Alexandra A. J. Migchielsen Yong Li Yu

Abstract A radioactive polymerase chain reaction (PCR) method has been developed for the relative quantification of the human α‐2 chain of type I collagen [hu α‐2 (I)] in cells. cDNAs generated by reverse transcription from the total pool of cytoplasmic RNA serve as a template for polymerase chain reaction amplification of a hu α‐2(I) cDNA primed by two sequence‐specific synthetic oligonucleotides. The distinctive 390 bp hu α‐2(I) cDNA and two Aval fragments of 220 and 170 bp are identified by agarose gel electrophoresis. α‐ 32 P‐dCTP of defined specific activity is included in the PCR reaction and the 390 bp cDNA is excised from the electrophoresis gel to permit direct radioactive quantification of hu α‐2(I) mRNA. The amount of hu α‐2(I) mRNA expressed in as few as 111 fibroblasts was determined reliably. In contrast, the hu α‐2(I) mRNA from at least 5 × 10 5 fibroblasts was required for detection by Northern blot analysis developed with the same cDNA probe radiolabelled with α‐ 32 P‐dCTP by random priming. Human bronchoalveolar lavage (BAL) fluids of six patients with fibrosing lung diseases stimulated the level of expression of hu α‐2(I) mRNA in cultured human fibroblasts as determined by this technique. The radioactive PCR method thus quantifies hu α‐2(I) mRNA in fibroblasts with sufficient sensitivity to study fibroblast activation in vitro and detect fibroblast stimuli in human clinical samples.

Agarose gel electrophoresis

Northern blot

10.1002/jcla.1860050407

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Citations (4)