Open AccessApplication of immunomagnetic beads in combination with RT‐PCR for the detection of circulating prostate cancer cells
Author(s) -
Makarovskiy Andrew N.,
Ackerley Wallace,
Wojcik Louis,
Halpert Gretchen K.,
Stein Barry S.,
Carreiro Marie P.,
Hixson Douglas C.
Publication year - 1997
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/(sici)1098-2825(1997)11:6<346::aid-jcla7>3.0.co;2-3
Subject(s) - prostate cancer , immunomagnetic separation , cancer detection , circulating tumor cell , cancer , computational biology , medicine , microbiology and biotechnology , biology , metastasis
Recently published protocols using Reverse Transcriptase Polymerase Chain reaction (RT‐PCR) for prostate specific antigen (PSA) provide a sensitive means for detecting circulating prostate cancer cells. Attempts to use these assays for staging of prostate cancer have produced conflicting results. As a first step towards rectifying these discrepancies, a modified immunobead‐RT‐PCR assay capable of detecting as few as 10 prostate cancer cells in 8cc of blood was developed. This 10 fold increase in sensitivity was achieved in part by introducing two target cell enrichment steps. As a model system to assess sensitivity of the modified assay, template RNA was extracted from PSA positive human carcinoma cells suspended in human blood and isolated with immunomagnetic beads following incubation with an epithelium specific antibody. After 45 cycles of PCR, product from as few as 10 target cells could be readily detected when displayed on a 2% agarose gel stained with SYBR Green fluorescent dye. The identity of amplified DNA fragments was confirmed by Southern blot hybridization. When applied to blood samples from patients with proven metastatic disease, the immuno‐bead RT‐PCR assay was successful in detecting circulating PSA positive epithelial cells, suggesting this assay may be useful for assessment of disease progression or recurrence. J. Clin. Lab. Anal. 11:346–350, 1997. © 1997 Wiley‐Liss, Inc.