The use of flow cytometry to examine calcium signalling by TRPV1 in mixed cell populations
Flow cytometric analysis of calcium mobilisation has been in use for many years in the study of specific receptor engagement or isolated cell:cell communication. However, calcium mobilisation/signaling is key to many cell functions including apoptosis, mobility and immune responses. Here we combine...
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| Veröffentlicht in: | Analytical biochemistry 2017-06, Vol.527, p.13-19 |
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| container_title | Analytical biochemistry |
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| creator | Assas, Bakri M. Abdulaal, Wesam H. Wakid, Majed H. Zakai, Haytham A. Miyan, J. Pennock, J.L. |
| description | Flow cytometric analysis of calcium mobilisation has been in use for many years in the study of specific receptor engagement or isolated cell:cell communication. However, calcium mobilisation/signaling is key to many cell functions including apoptosis, mobility and immune responses. Here we combine multiplex surface staining of whole spleen with Indo-1 AM to visualise calcium mobilisation and examine calcium signaling in a mixed immune cell culture over time. We demonstrate responses to a TRPV1 agonist in distinct cell subtypes without the need for cell separation. Multi parameter staining alongside Indo-1 AM to demonstrate calcium mobilization allows the study of real time calcium signaling in a complex environment.
•This paper aims to help discuss calcium signaling in TRPV1+ splenocytes via flow cytomerty.•We re-introduce a previously used technique to help identify different cell populations accurately with adequate sensitivity.•This technique is economical and easy to reproduce. |
| doi_str_mv | 10.1016/j.ab.2017.03.025 |
| format | Article |
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•This paper aims to help discuss calcium signaling in TRPV1+ splenocytes via flow cytomerty.•We re-introduce a previously used technique to help identify different cell populations accurately with adequate sensitivity.•This technique is economical and easy to reproduce.</description><identifier>ISSN: 0003-2697</identifier><identifier>EISSN: 1096-0309</identifier><identifier>DOI: 10.1016/j.ab.2017.03.025</identifier><identifier>PMID: 28372979</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Animals ; Antigens, CD - genetics ; Antigens, CD - immunology ; B-Lymphocytes - cytology ; B-Lymphocytes - drug effects ; B-Lymphocytes - immunology ; Biomarkers ; Calcium - immunology ; Calcium - metabolism ; Calcium Signaling ; Capsaicin - pharmacology ; Dendritic Cells - cytology ; Dendritic Cells - drug effects ; Dendritic Cells - immunology ; Flow cytometry ; Flow Cytometry - methods ; Gene Expression ; Immune phenotyping ; Indo-1 AM ; Indoles - chemistry ; Macrophages - cytology ; Macrophages - drug effects ; Macrophages - immunology ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Primary Cell Culture ; Spleen - cytology ; Spleen - immunology ; Staining and Labeling - methods ; T-Lymphocytes - cytology ; T-Lymphocytes - drug effects ; T-Lymphocytes - immunology ; TRPV Cation Channels - deficiency ; TRPV Cation Channels - genetics ; TRPV Cation Channels - immunology ; TRPV1</subject><ispartof>Analytical biochemistry, 2017-06, Vol.527, p.13-19</ispartof><rights>2017</rights><rights>Copyright © 2017. Published by Elsevier Inc.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c350t-1d3af1ec6e4b4906dfca5c12799537ef59211e92d97635ce9233a7fc1287b09a3</citedby><cites>FETCH-LOGICAL-c350t-1d3af1ec6e4b4906dfca5c12799537ef59211e92d97635ce9233a7fc1287b09a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.ab.2017.03.025$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,777,781,3537,27905,27906,45976</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/28372979$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Assas, Bakri M.</creatorcontrib><creatorcontrib>Abdulaal, Wesam H.</creatorcontrib><creatorcontrib>Wakid, Majed H.</creatorcontrib><creatorcontrib>Zakai, Haytham A.</creatorcontrib><creatorcontrib>Miyan, J.</creatorcontrib><creatorcontrib>Pennock, J.L.</creatorcontrib><title>The use of flow cytometry to examine calcium signalling by TRPV1 in mixed cell populations</title><title>Analytical biochemistry</title><addtitle>Anal Biochem</addtitle><description>Flow cytometric analysis of calcium mobilisation has been in use for many years in the study of specific receptor engagement or isolated cell:cell communication. However, calcium mobilisation/signaling is key to many cell functions including apoptosis, mobility and immune responses. Here we combine multiplex surface staining of whole spleen with Indo-1 AM to visualise calcium mobilisation and examine calcium signaling in a mixed immune cell culture over time. We demonstrate responses to a TRPV1 agonist in distinct cell subtypes without the need for cell separation. Multi parameter staining alongside Indo-1 AM to demonstrate calcium mobilization allows the study of real time calcium signaling in a complex environment.
•This paper aims to help discuss calcium signaling in TRPV1+ splenocytes via flow cytomerty.•We re-introduce a previously used technique to help identify different cell populations accurately with adequate sensitivity.•This technique is economical and easy to reproduce.</description><subject>Animals</subject><subject>Antigens, CD - genetics</subject><subject>Antigens, CD - immunology</subject><subject>B-Lymphocytes - cytology</subject><subject>B-Lymphocytes - drug effects</subject><subject>B-Lymphocytes - immunology</subject><subject>Biomarkers</subject><subject>Calcium - immunology</subject><subject>Calcium - metabolism</subject><subject>Calcium Signaling</subject><subject>Capsaicin - pharmacology</subject><subject>Dendritic Cells - cytology</subject><subject>Dendritic Cells - drug effects</subject><subject>Dendritic Cells - immunology</subject><subject>Flow cytometry</subject><subject>Flow Cytometry - methods</subject><subject>Gene Expression</subject><subject>Immune phenotyping</subject><subject>Indo-1 AM</subject><subject>Indoles - chemistry</subject><subject>Macrophages - cytology</subject><subject>Macrophages - drug effects</subject><subject>Macrophages - immunology</subject><subject>Male</subject><subject>Mice</subject><subject>Mice, Inbred C57BL</subject><subject>Mice, Knockout</subject><subject>Primary Cell Culture</subject><subject>Spleen - cytology</subject><subject>Spleen - immunology</subject><subject>Staining and Labeling - methods</subject><subject>T-Lymphocytes - cytology</subject><subject>T-Lymphocytes - drug effects</subject><subject>T-Lymphocytes - immunology</subject><subject>TRPV Cation Channels - deficiency</subject><subject>TRPV Cation Channels - genetics</subject><subject>TRPV Cation Channels - immunology</subject><subject>TRPV1</subject><issn>0003-2697</issn><issn>1096-0309</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kE1P4zAQhi20iJaPO6eVj3tJmLGbuN7bCvElIYFQ4cDFcpxJ11USd-ME2n9PqsLeOM0cnvfVzMPYOUKKgPnFKrVFKgBVCjIFkR2wKYLOE5Cgf7ApAMhE5FpN2HGMKwDEWZYfsYmYSyW00lP2uvhLfIjEQ8WrOrxzt-1DQ3235X3gtLGNb4k7Wzs_NDz6ZWvr2rdLXmz54unxBblveeM3VHJHdc3XYT3UtvehjafssLJ1pLPPecKer68Wl7fJ_cPN3eWf-8TJDPoES2krJJfTrJhpyMvK2cyhUFpnUlGVaYFIWpRa5TJz4yalVdVIzFUB2soT9mvfu-7Cv4Fibxofd8fYlsIQDc7nM8wzgXpEYY-6LsTYUWXWnW9stzUIZmfUrIwtzM6oAWlGo2Pk52f7UDRU_g98KRyB33uAxh_fPHUmOk-to9J35HpTBv99-wf-Q4UR</recordid><startdate>20170615</startdate><enddate>20170615</enddate><creator>Assas, Bakri M.</creator><creator>Abdulaal, Wesam H.</creator><creator>Wakid, Majed H.</creator><creator>Zakai, Haytham A.</creator><creator>Miyan, J.</creator><creator>Pennock, J.L.</creator><general>Elsevier Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20170615</creationdate><title>The use of flow cytometry to examine calcium signalling by TRPV1 in mixed cell populations</title><author>Assas, Bakri M. ; Abdulaal, Wesam H. ; Wakid, Majed H. ; Zakai, Haytham A. ; Miyan, J. ; Pennock, J.L.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c350t-1d3af1ec6e4b4906dfca5c12799537ef59211e92d97635ce9233a7fc1287b09a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Animals</topic><topic>Antigens, CD - genetics</topic><topic>Antigens, CD - immunology</topic><topic>B-Lymphocytes - cytology</topic><topic>B-Lymphocytes - drug effects</topic><topic>B-Lymphocytes - immunology</topic><topic>Biomarkers</topic><topic>Calcium - immunology</topic><topic>Calcium - metabolism</topic><topic>Calcium Signaling</topic><topic>Capsaicin - pharmacology</topic><topic>Dendritic Cells - cytology</topic><topic>Dendritic Cells - drug effects</topic><topic>Dendritic Cells - immunology</topic><topic>Flow cytometry</topic><topic>Flow Cytometry - methods</topic><topic>Gene Expression</topic><topic>Immune phenotyping</topic><topic>Indo-1 AM</topic><topic>Indoles - chemistry</topic><topic>Macrophages - cytology</topic><topic>Macrophages - drug effects</topic><topic>Macrophages - immunology</topic><topic>Male</topic><topic>Mice</topic><topic>Mice, Inbred C57BL</topic><topic>Mice, Knockout</topic><topic>Primary Cell Culture</topic><topic>Spleen - cytology</topic><topic>Spleen - immunology</topic><topic>Staining and Labeling - methods</topic><topic>T-Lymphocytes - cytology</topic><topic>T-Lymphocytes - drug effects</topic><topic>T-Lymphocytes - immunology</topic><topic>TRPV Cation Channels - deficiency</topic><topic>TRPV Cation Channels - genetics</topic><topic>TRPV Cation Channels - immunology</topic><topic>TRPV1</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Assas, Bakri M.</creatorcontrib><creatorcontrib>Abdulaal, Wesam H.</creatorcontrib><creatorcontrib>Wakid, Majed H.</creatorcontrib><creatorcontrib>Zakai, Haytham A.</creatorcontrib><creatorcontrib>Miyan, J.</creatorcontrib><creatorcontrib>Pennock, J.L.</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Analytical biochemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Assas, Bakri M.</au><au>Abdulaal, Wesam H.</au><au>Wakid, Majed H.</au><au>Zakai, Haytham A.</au><au>Miyan, J.</au><au>Pennock, J.L.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>The use of flow cytometry to examine calcium signalling by TRPV1 in mixed cell populations</atitle><jtitle>Analytical biochemistry</jtitle><addtitle>Anal Biochem</addtitle><date>2017-06-15</date><risdate>2017</risdate><volume>527</volume><spage>13</spage><epage>19</epage><pages>13-19</pages><issn>0003-2697</issn><eissn>1096-0309</eissn><abstract>Flow cytometric analysis of calcium mobilisation has been in use for many years in the study of specific receptor engagement or isolated cell:cell communication. However, calcium mobilisation/signaling is key to many cell functions including apoptosis, mobility and immune responses. Here we combine multiplex surface staining of whole spleen with Indo-1 AM to visualise calcium mobilisation and examine calcium signaling in a mixed immune cell culture over time. We demonstrate responses to a TRPV1 agonist in distinct cell subtypes without the need for cell separation. Multi parameter staining alongside Indo-1 AM to demonstrate calcium mobilization allows the study of real time calcium signaling in a complex environment.
•This paper aims to help discuss calcium signaling in TRPV1+ splenocytes via flow cytomerty.•We re-introduce a previously used technique to help identify different cell populations accurately with adequate sensitivity.•This technique is economical and easy to reproduce.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>28372979</pmid><doi>10.1016/j.ab.2017.03.025</doi><tpages>7</tpages></addata></record> |
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| subjects | Animals Antigens, CD - genetics Antigens, CD - immunology B-Lymphocytes - cytology B-Lymphocytes - drug effects B-Lymphocytes - immunology Biomarkers Calcium - immunology Calcium - metabolism Calcium Signaling Capsaicin - pharmacology Dendritic Cells - cytology Dendritic Cells - drug effects Dendritic Cells - immunology Flow cytometry Flow Cytometry - methods Gene Expression Immune phenotyping Indo-1 AM Indoles - chemistry Macrophages - cytology Macrophages - drug effects Macrophages - immunology Male Mice Mice, Inbred C57BL Mice, Knockout Primary Cell Culture Spleen - cytology Spleen - immunology Staining and Labeling - methods T-Lymphocytes - cytology T-Lymphocytes - drug effects T-Lymphocytes - immunology TRPV Cation Channels - deficiency TRPV Cation Channels - genetics TRPV Cation Channels - immunology TRPV1 |
| title | The use of flow cytometry to examine calcium signalling by TRPV1 in mixed cell populations |
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