TY - JOUR
T1 - Detection of tumor-associated cells in cryopreserved peripheral blood mononuclear cell samples for retrospective analysis
AU - Zhu, Peixuan
AU - Stanton, Melissa L.
AU - Castle, Erik P.
AU - Joseph, Richard W.
AU - Adams, Daniel L.
AU - Li, Shuhong
AU - Amstutz, Platte
AU - Tang, Cha Mei
AU - Ho, Thai H.
N1 - Funding Information:
The authors acknowledge the support provided by the Gloria A. and Thomas J. Dutson Jr, Kidney Research Endowment. This work was supported by the Mayo Clinic Center for Individualized Medicine (T.H.H.); a Gerstner Family Career Development Award (T.H.H., R.W.J.); Kathryn H. and Roger Penske Career Development Award to Support Medical Research (T.H.H.); the National Cancer Institute at the National Institutes of Health (NIH) (K12CA90628) (T.H.H.). The US Army Research Office (ARO) and the Defense Advanced Research Projects Agency (DARPA) (W911NF‑14‑C‑0098) (C.‑M.T.). This publica‑ tion was made possible by Clinical and Translational Science Awards Grant Number UL1 TR000135 from the National Center for Advancing Translational Sciences (NCATS), a component of the NIH. The funders had no role in the design of the study and collection, analysis, interpretation of data or in writing the manuscript.
Funding Information:
The authors acknowledge the support provided by the Gloria A. and Thomas J. Dutson Jr., Kidney Research Endowment. This work was supported by the Mayo Clinic Center for Individualized Medicine (T.H.H.); a Gerstner Family Career Development Award (T.H.H., R.W.J.); Kathryn H. and Roger Penske Career Development Award to Support Medical Research (T.H.H.); the National Cancer Institute at the National Institutes of Health (NIH) (K12CA90628) (T.H.H.). The US Army Research Office (ARO) and the Defense Advanced Research Projects Agency (DARPA) (W911NF‑14‑C‑0098) (C.‑M.T.). This publica‑ tion was made possible by Clinical and Translational Science Awards Grant Number UL1 TR000135 from the National Center for Advancing Translational Sciences (NCATS), a component of the NIH. Its contents are solely the respon‑ sibility of the authors and do not necessarily represent the official views of the NIH or the policy of the US Government.
Publisher Copyright:
© 2016 The Author(s).
PY - 2016/7/2
Y1 - 2016/7/2
N2 - Background: Cryopreserved peripheral blood mononuclear cells (PBMCs) are commonly collected in biobanks. However, little data exist regarding the preservation of tumor-associated cells in cryopreserved collections. The objective of this study was to determine the feasibility of using the CellSieve™ microfiltration assay for the isolation of circulating tumor cells (CTCs) and circulating cancer-associated macrophage-like cells (CAMLs) from cryopreserved PBMC samples. Methods: Blood samples spiked with breast (MCF-7), prostate (PC-3), and renal (786-O) cancer cell lines were used to establish analytical accuracy, efficiency, and reproducibility after cryopreservation. The spiked samples were processed through Ficoll separation, and cryopreservation was followed by thawing and microfiltration. Results: MCF-7 cells were successfully retrieved with recovery efficiencies of 90.5 % without cryopreservation and 87.8 and 89.0 %, respectively, on day 7 and day 66 following cryopreservation. The corresponding recovery efficiencies of PC-3 cells were 83.3 % without cryopreservation and 85.3 and 84.7 %, respectively, after cryopreservation. Recovery efficiencies of 786-O cells were 92.7 % without cryopreservation, and 82.7 and 81.3 %, respectively, after cryopreservation. The recovered cells retained the morphologic characteristics and immunohistochemical markers that had been observed before freezing. The protocols were further validated by quantitation of CAMLs in blood samples from two patients with renal cell carcinoma (RCC). The recovery rates of CTCs and CAMLs from cryopreserved samples were not statistically significant different (P > 0.05) from matched fresh samples. Conclusions: To our knowledge, this is the first report that CAMLs could be cryopreserved and analyzed after thawing with microfiltration technology. The application of microfiltration technology to cryopreserved samples will enable much greater retrospective study of cancer patients in relation to long-term outcomes.
AB - Background: Cryopreserved peripheral blood mononuclear cells (PBMCs) are commonly collected in biobanks. However, little data exist regarding the preservation of tumor-associated cells in cryopreserved collections. The objective of this study was to determine the feasibility of using the CellSieve™ microfiltration assay for the isolation of circulating tumor cells (CTCs) and circulating cancer-associated macrophage-like cells (CAMLs) from cryopreserved PBMC samples. Methods: Blood samples spiked with breast (MCF-7), prostate (PC-3), and renal (786-O) cancer cell lines were used to establish analytical accuracy, efficiency, and reproducibility after cryopreservation. The spiked samples were processed through Ficoll separation, and cryopreservation was followed by thawing and microfiltration. Results: MCF-7 cells were successfully retrieved with recovery efficiencies of 90.5 % without cryopreservation and 87.8 and 89.0 %, respectively, on day 7 and day 66 following cryopreservation. The corresponding recovery efficiencies of PC-3 cells were 83.3 % without cryopreservation and 85.3 and 84.7 %, respectively, after cryopreservation. Recovery efficiencies of 786-O cells were 92.7 % without cryopreservation, and 82.7 and 81.3 %, respectively, after cryopreservation. The recovered cells retained the morphologic characteristics and immunohistochemical markers that had been observed before freezing. The protocols were further validated by quantitation of CAMLs in blood samples from two patients with renal cell carcinoma (RCC). The recovery rates of CTCs and CAMLs from cryopreserved samples were not statistically significant different (P > 0.05) from matched fresh samples. Conclusions: To our knowledge, this is the first report that CAMLs could be cryopreserved and analyzed after thawing with microfiltration technology. The application of microfiltration technology to cryopreserved samples will enable much greater retrospective study of cancer patients in relation to long-term outcomes.
KW - CellSieve™ microfiltration
KW - Circulating cancer-associated macrophage-like cells (CAMLs)
KW - Circulating tumor cells (CTCs)
KW - Cryopreservation
KW - Ficoll
KW - Fluorescence antibody staining
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U2 - 10.1186/s12967-016-0953-2
DO - 10.1186/s12967-016-0953-2
M3 - Article
C2 - 27369977
AN - SCOPUS:84976863794
SN - 1479-5876
VL - 14
JO - Journal of Translational Medicine
JF - Journal of Translational Medicine
IS - 1
M1 - 198
ER -