A collection of primary tissue cultures of tumors from vacuum packed and cooled surgical specimens: a feasibility study
Primary cultures represent an invaluable tool to set up functional experimental conditions; however, creation of tissue cultures from solid tumors is troublesome and often unproductive. Several features can affect the success rate of primary cultures, including technical issues from pre-analytical p...
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| Veröffentlicht in: | PloS one 2013-09, Vol.8 (9), p.e75193-e75193 |
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| creator | Annaratone, Laura Marchiò, Caterina Russo, Rosalia Ciardo, Luigi Rondon-Lagos, Sandra Milena Goia, Margherita Scalzo, Maria Stella Bolla, Stefania Castellano, Isabella Verdun di Cantogno, Ludovica Bussolati, Gianni Sapino, Anna |
| description | Primary cultures represent an invaluable tool to set up functional experimental conditions; however, creation of tissue cultures from solid tumors is troublesome and often unproductive. Several features can affect the success rate of primary cultures, including technical issues from pre-analytical procedures employed in surgical theaters and pathology laboratories. We have recently introduced a new method of collection, transfer, and preservation of surgical specimens that requires immediate vacuum sealing of excised specimens at surgical theaters, followed by time-controlled transferring at 4°C to the pathology laboratory. Here we investigate the feasibility and performance of short-term primary cell cultures derived from vacuum packed and cooled (VPAC) preserved tissues. Tissue fragments were sampled from 52 surgical specimens of tumors larger than 2 cm for which surgical and VPAC times (the latter corresponding to cold ischemia time) were recorded. Cell viability was determined by trypan blue dye-exclusion assay and hematoxylin and eosin and immunohistochemical stainings were performed to appreciate morphological and immunophenotypical features of cultured cells. Cell viability showed a range of 84-100% in 44 out of 52 (85%) VPAC preserved tissues. Length of both surgical and VPAC times affected cell viability: the critical surgical time was set around 1 hour and 30 minutes, while cells preserved a good viability when kept for about 24 hours of vacuum at 4°C. Cells were maintained in culture for at least three passages. Immunocytochemistry confirmed the phenotype of distinct populations, that is, expression of cytokeratins in epithelioid cells and of vimentin in spindle cells. Our results suggest that VPAC preserved tissues may represent a reliable source for creation of primary cell cultures and that a careful monitoring of surgical and cold ischemia times fosters a good performance of primary tissue cultures. |
| doi_str_mv | 10.1371/journal.pone.0075193 |
| format | Article |
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Several features can affect the success rate of primary cultures, including technical issues from pre-analytical procedures employed in surgical theaters and pathology laboratories. We have recently introduced a new method of collection, transfer, and preservation of surgical specimens that requires immediate vacuum sealing of excised specimens at surgical theaters, followed by time-controlled transferring at 4°C to the pathology laboratory. Here we investigate the feasibility and performance of short-term primary cell cultures derived from vacuum packed and cooled (VPAC) preserved tissues. Tissue fragments were sampled from 52 surgical specimens of tumors larger than 2 cm for which surgical and VPAC times (the latter corresponding to cold ischemia time) were recorded. Cell viability was determined by trypan blue dye-exclusion assay and hematoxylin and eosin and immunohistochemical stainings were performed to appreciate morphological and immunophenotypical features of cultured cells. Cell viability showed a range of 84-100% in 44 out of 52 (85%) VPAC preserved tissues. Length of both surgical and VPAC times affected cell viability: the critical surgical time was set around 1 hour and 30 minutes, while cells preserved a good viability when kept for about 24 hours of vacuum at 4°C. Cells were maintained in culture for at least three passages. Immunocytochemistry confirmed the phenotype of distinct populations, that is, expression of cytokeratins in epithelioid cells and of vimentin in spindle cells. Our results suggest that VPAC preserved tissues may represent a reliable source for creation of primary cell cultures and that a careful monitoring of surgical and cold ischemia times fosters a good performance of primary tissue cultures.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0075193</identifier><identifier>PMID: 24098684</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Biotechnology ; Breast cancer ; Cell culture ; Cold ; Cold Temperature ; Collection ; Epidermal growth factor ; Estrogens ; Feasibility Studies ; Gene expression ; Hospitals ; Humans ; Immunocytochemistry ; Ischemia ; Laboratories ; Medicine ; Neoplasms - surgery ; Pathology ; Penicillin ; Preservation ; Solid tumors ; Specimen Handling - methods ; Surgery ; Surgical instruments ; Theaters & cinemas ; Tissue Culture Techniques - methods ; Tissue Preservation - methods ; Tissues ; Transplants & implants ; Tumors ; Vacuum ; Vimentin</subject><ispartof>PloS one, 2013-09, Vol.8 (9), p.e75193-e75193</ispartof><rights>2013 Annaratone et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License: https://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 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Several features can affect the success rate of primary cultures, including technical issues from pre-analytical procedures employed in surgical theaters and pathology laboratories. We have recently introduced a new method of collection, transfer, and preservation of surgical specimens that requires immediate vacuum sealing of excised specimens at surgical theaters, followed by time-controlled transferring at 4°C to the pathology laboratory. Here we investigate the feasibility and performance of short-term primary cell cultures derived from vacuum packed and cooled (VPAC) preserved tissues. Tissue fragments were sampled from 52 surgical specimens of tumors larger than 2 cm for which surgical and VPAC times (the latter corresponding to cold ischemia time) were recorded. Cell viability was determined by trypan blue dye-exclusion assay and hematoxylin and eosin and immunohistochemical stainings were performed to appreciate morphological and immunophenotypical features of cultured cells. Cell viability showed a range of 84-100% in 44 out of 52 (85%) VPAC preserved tissues. Length of both surgical and VPAC times affected cell viability: the critical surgical time was set around 1 hour and 30 minutes, while cells preserved a good viability when kept for about 24 hours of vacuum at 4°C. Cells were maintained in culture for at least three passages. Immunocytochemistry confirmed the phenotype of distinct populations, that is, expression of cytokeratins in epithelioid cells and of vimentin in spindle cells. Our results suggest that VPAC preserved tissues may represent a reliable source for creation of primary cell cultures and that a careful monitoring of surgical and cold ischemia times fosters a good performance of primary tissue cultures.</description><subject>Biotechnology</subject><subject>Breast cancer</subject><subject>Cell culture</subject><subject>Cold</subject><subject>Cold Temperature</subject><subject>Collection</subject><subject>Epidermal growth factor</subject><subject>Estrogens</subject><subject>Feasibility Studies</subject><subject>Gene expression</subject><subject>Hospitals</subject><subject>Humans</subject><subject>Immunocytochemistry</subject><subject>Ischemia</subject><subject>Laboratories</subject><subject>Medicine</subject><subject>Neoplasms - surgery</subject><subject>Pathology</subject><subject>Penicillin</subject><subject>Preservation</subject><subject>Solid tumors</subject><subject>Specimen Handling - methods</subject><subject>Surgery</subject><subject>Surgical instruments</subject><subject>Theaters & cinemas</subject><subject>Tissue Culture Techniques - 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Annaratone, Laura</au><au>Marchiò, Caterina</au><au>Russo, Rosalia</au><au>Ciardo, Luigi</au><au>Rondon-Lagos, Sandra Milena</au><au>Goia, Margherita</au><au>Scalzo, Maria Stella</au><au>Bolla, Stefania</au><au>Castellano, Isabella</au><au>Verdun di Cantogno, Ludovica</au><au>Bussolati, Gianni</au><au>Sapino, Anna</au><au>Burns, Jorge Sans</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A collection of primary tissue cultures of tumors from vacuum packed and cooled surgical specimens: a feasibility study</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2013-09-30</date><risdate>2013</risdate><volume>8</volume><issue>9</issue><spage>e75193</spage><epage>e75193</epage><pages>e75193-e75193</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Primary cultures represent an invaluable tool to set up functional experimental conditions; however, creation of tissue cultures from solid tumors is troublesome and often unproductive. Several features can affect the success rate of primary cultures, including technical issues from pre-analytical procedures employed in surgical theaters and pathology laboratories. We have recently introduced a new method of collection, transfer, and preservation of surgical specimens that requires immediate vacuum sealing of excised specimens at surgical theaters, followed by time-controlled transferring at 4°C to the pathology laboratory. Here we investigate the feasibility and performance of short-term primary cell cultures derived from vacuum packed and cooled (VPAC) preserved tissues. Tissue fragments were sampled from 52 surgical specimens of tumors larger than 2 cm for which surgical and VPAC times (the latter corresponding to cold ischemia time) were recorded. Cell viability was determined by trypan blue dye-exclusion assay and hematoxylin and eosin and immunohistochemical stainings were performed to appreciate morphological and immunophenotypical features of cultured cells. Cell viability showed a range of 84-100% in 44 out of 52 (85%) VPAC preserved tissues. Length of both surgical and VPAC times affected cell viability: the critical surgical time was set around 1 hour and 30 minutes, while cells preserved a good viability when kept for about 24 hours of vacuum at 4°C. Cells were maintained in culture for at least three passages. Immunocytochemistry confirmed the phenotype of distinct populations, that is, expression of cytokeratins in epithelioid cells and of vimentin in spindle cells. Our results suggest that VPAC preserved tissues may represent a reliable source for creation of primary cell cultures and that a careful monitoring of surgical and cold ischemia times fosters a good performance of primary tissue cultures.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>24098684</pmid><doi>10.1371/journal.pone.0075193</doi><oa>free_for_read</oa></addata></record> |
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| subjects | Biotechnology Breast cancer Cell culture Cold Cold Temperature Collection Epidermal growth factor Estrogens Feasibility Studies Gene expression Hospitals Humans Immunocytochemistry Ischemia Laboratories Medicine Neoplasms - surgery Pathology Penicillin Preservation Solid tumors Specimen Handling - methods Surgery Surgical instruments Theaters & cinemas Tissue Culture Techniques - methods Tissue Preservation - methods Tissues Transplants & implants Tumors Vacuum Vimentin |
| title | A collection of primary tissue cultures of tumors from vacuum packed and cooled surgical specimens: a feasibility study |
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