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{{About|Carboxyfluorescein succinimidyl ester|Crystal field stabilization energy|Crystal field stabilization energy}} {{About|Carboxyfluorescein succinimidyl ester|Crystal field stabilization energy|Crystal field stabilization energy}}

{{Chembox {{Chembox
| Verifiedfields = changed
⚫ | verifiedrevid = 413510358
| Watchedfields = changed
⚫ | verifiedrevid = 439546511
| Name=6-Carboxyfluorescein succinimidyl ester | Name=6-Carboxyfluorescein succinimidyl ester
| ImageFile = Carboxyfluorescein succinimidyl ester.png | ImageFile = Carboxyfluorescein succinimidyl ester.png
| ImageSize = 200px | ImageSize =
| PIN = 2,5-Dioxopyrrolidin-1-yl 3,6-dihydroxy-3-oxo-3''H''-spirobenzofuran-1,9′-xanthene]-6-carboxylate
| IUPACName =
| OtherNames = CFSE; Carboxyfluorescein ''N''-succinimidyl ester | OtherNames = CFSE; Carboxyfluorescein ''N''-succinimidyl ester
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| CASOther = <br>117548-22-8 (5/6 mixture)
| CASNo2 = 117548-22-8
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| InChI = 1/C25H15NO9/c27-13-2-5-16-19(10-13)33-20-11-14(28)3-6-17(20)25(16)18-9-12(1-4-15(18)24(32)34-25)23(31)35-26-21(29)7-8-22(26)30/h1-6,9-11,27-28H,7-8H2
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'''Carboxyfluorescein succinimidyl ester''' (CFSE) is a ] cell ] dye. CFSE is commonly confused with ] (CFDA-SE), although they are not strictly the same molecule; CFDA-SE, due to its acetate groups, is highly cell permeable, while CFSE is much less so. As CFDA-SE, which is non-fluorescent, enters the cytoplasm of cells, intracellular esterases remove the acetate groups and convert the molecule to the fluorescent ester, CFSE, which is retained within cells and covalently couples, via its succinimidyl group, to intracellular molecules.<ref>{{cite journal |author=Parish CR |title=Fluorescent dyes for lymphocyte migration and proliferation studies |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=499–508 |year=1999 |month=December |pmid=10571670 |doi=10.1046/j.1440-1711.1999.00877.x }}</ref> Due to this covalent coupling reaction fluorescent CFSE can be retained within cells for extremely long periods. Also, due to this stable linkage, once incorporated within cells the dye is not transferred to adjacent cells. '''Carboxyfluorescein succinimidyl ester''' ('''CFSE''') is a ] cell ] dye. CFSE is cell permeable and covalently couples, via its ], to intracellular molecules,<ref>{{cite journal |author=Parish CR |title=Fluorescent dyes for lymphocyte migration and proliferation studies |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=499–508 |date=December 1999 |pmid=10571670 |doi=10.1046/j.1440-1711.1999.00877.x |s2cid=2194612 }}</ref> notably, to intracellular ] residues and other amine sources. Due to this covalent coupling reaction, fluorescent CFSE can be retained within cells for extremely long periods. Also, due to this stable linkage, once incorporated within cells, the dye is not transferred to adjacent cells.

CFSE is commonly confused with ] (CFDA-SE), although they are not strictly the same molecule; CFDA-SE, due to its acetate groups, is highly cell permeable, while CFSE is much less so. As CFDA-SE, which is non-fluorescent, enters the ] of ], intracellular esterases remove the acetate groups and convert the molecule to the fluorescent ester.


CFSE was originally developed as a fluorescent dye that could be used to stably label lymphocytes and track their migration within animals for many months.<ref>{{cite journal |author=Weston SA, Parish CR |title=New fluorescent dyes for lymphocyte migration studies. Analysis by flow cytometry and fluorescence microscopy |journal=Journal of Immunological Methods |volume=133 |issue=1 |pages=87–97 |year=1990 |month=October |pmid=2212694 |doi= 10.1016/0022-1759(90)90322-M|url=http://linkinghub.elsevier.com/retrieve/pii/0022-1759(90)90322-M}}</ref> Subsequent studies revealed that the dye can be used to monitor lymphocyte proliferation, both ''in vitro'' and ''in vivo'', due to the progressive halving of CFSE fluorescence within daughter cells following each cell division.<ref>{{cite journal |author=Lyons AB, Parish CR |title=Determination of lymphocyte division by flow cytometry |journal=Journal of Immunological Methods |volume=171 |issue=1 |pages=131–7 |year=1994 |month=May |pmid=8176234 |doi= 10.1016/0022-1759(94)90236-4|url=http://linkinghub.elsevier.com/retrieve/pii/0022-1759(94)90236-4}}</ref> The only limitation is that CFSE at high concentrations can be toxic for cells. However, when CFSE labelling is performed optimally, approximately 7-8 cell divisions can be identified before the CFSE fluorescence is too low to be distinguished above the autofluorescence background. Thus CFSE represents an extremely valuable fluorescent dye for immunological studies, allowing lymphocyte proliferation, migration and positioning to be simultaneously monitored. By the use of fluorescent antibodies against different lymphocyte cell surface markers it is also possible to follow the proliferation behaviour of different lymphocyte subsets.<ref>{{cite journal |author=Fazekas de St Groth B, Smith AL, Koh WP, Girgis L, Cook MC, Bertolino P |title=Carboxyfluorescein diacetate succinimidyl ester and the virgin lymphocyte: a marriage made in heaven |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=530–8 |year=1999 |month=December |pmid=10571674 |doi=10.1046/j.1440-1711.1999.00871.x }}</ref> In addition, unlike other methods, CFSE-labeled viable cells can be recovered for further analysis. CFSE was originally developed as a fluorescent dye that could be used to stably label ]s and track their migration within animals for many months.<ref>{{cite journal |vauthors=Weston SA, Parish CR |title=New fluorescent dyes for lymphocyte migration studies. Analysis by flow cytometry and fluorescence microscopy |journal=Journal of Immunological Methods |volume=133 |issue=1 |pages=87–97 |date=October 1990 |pmid=2212694 |doi= 10.1016/0022-1759(90)90322-M}}</ref> Subsequent studies revealed that the dye can be used to monitor lymphocyte ], both ''in vitro'' and ''in vivo'', due to the progressive halving of CFSE fluorescence within daughter cells following each cell division.<ref>{{cite journal |vauthors=Lyons AB, Parish CR |title=Determination of lymphocyte division by flow cytometry |journal=Journal of Immunological Methods |volume=171 |issue=1 |pages=131–7 |date=May 1994 |pmid=8176234 |doi= 10.1016/0022-1759(94)90236-4}}</ref> The only limitation is that CFSE at high concentrations can be toxic for cells. However, when CFSE labelling is performed optimally, approximately 7-8 cell divisions can be identified before the CFSE fluorescence is too low to be distinguished above the autofluorescence background. Thus CFSE represents an extremely valuable fluorescent dye for immunological studies, allowing lymphocyte proliferation, migration and positioning to be simultaneously monitored. By the use of fluorescent antibodies against different lymphocyte cell surface markers it is also possible to follow the proliferation behaviour of different lymphocyte subsets.<ref>{{cite journal |vauthors=Fazekas de St Groth B, Smith AL, Koh WP, Girgis L, Cook MC, Bertolino P |title=Carboxyfluorescein diacetate succinimidyl ester and the virgin lymphocyte: a marriage made in heaven |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=530–8 |date=December 1999 |pmid=10571674 |doi=10.1046/j.1440-1711.1999.00871.x |s2cid=2786618 }}</ref> In addition, unlike other methods, CFSE-labeled viable cells can be recovered for further analysis.


Since the initial description of CFSE it has been used in thousands of immunological studies, an example of an early proliferation study in animals being described by Kurts et.al.<ref>{{cite journal |author=Kurts C, Kosaka H, Carbone FR, Miller JF, Heath WR |title=Class I-restricted cross-presentation of exogenous self-antigens leads to deletion of autoreactive CD8(+) T cells |journal=The Journal of Experimental Medicine |volume=186 |issue=2 |pages=239–45 |year=1997 |month=July |pmid=9221753 |pmc=2198972 |doi= |url=http://www.jem.org/cgi/pmidlookup?view=long&pmid=9221753}}</ref> However, perhaps the most important CFSE investigations have been those demonstrating that many of the effector functions of lymphocytes, such as cytokine production by T lymphocytes,<ref>{{cite journal |author=Gett AV, Hodgkin PD |title=Cell division regulates the T cell cytokine repertoire, revealing a mechanism underlying immune class regulation |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=95 |issue=16 |pages=9488–93 |year=1998 |month=August |pmid=9689107 |pmc=21365 |doi= |url=http://www.pnas.org/cgi/pmidlookup?view=long&pmid=9689107}}</ref><ref>{{cite journal |author=Bird JJ, Brown DR, Mullen AC, ''et al.'' |title=Helper T cell differentiation is controlled by the cell cycle |journal=Immunity |volume=9 |issue=2 |pages=229–37 |year=1998 |month=August |pmid=9729043 |doi= 10.1016/S1074-7613(00)80605-6|url=http://linkinghub.elsevier.com/retrieve/pii/S1074-7613(00)80605-6}}</ref> and antibody class switching by B cells,<ref>{{cite journal |author=Hodgkin PD, Lee JH, Lyons AB |title=B cell differentiation and isotype switching is related to division cycle number |journal=The Journal of Experimental Medicine |volume=184 |issue=1 |pages=277–81 |year=1996 |month=July |pmid=8691143 |pmc=2192686 |doi= |url=}}</ref> are division dependent. Sophisticated mathematical models have also been developed to analyse CFSE data and probe various aspects of immune responses.<ref>{{cite journal |author=Nordon RE, Nakamura M, Ramirez C, Odell R |title=Analysis of growth kinetics by division tracking |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=523–9 |year=1999 |month=December |pmid=10571673 |doi=10.1046/j.1440-1711.1999.00869.x }}</ref><ref>{{cite journal |author=Gett AV, Hodgkin PD |title=A cellular calculus for signal integration by T cells |journal=Nature Immunology |volume=1 |issue=3 |pages=239–44 |year=2000 |month=September |pmid=10973282 |doi=10.1038/79782 }}</ref><ref>{{cite journal |author=De Boer RJ, Ganusov VV, Milutinović D, Hodgkin PD, Perelson AS |title=Estimating lymphocyte division and death rates from CFSE data |journal=Bulletin of Mathematical Biology |volume=68 |issue=5 |pages=1011–31 |year=2006 |month=July |pmid=16832737 |doi=10.1007/s11538-006-9094-8 }}</ref><ref>{{cite journal |author=Callard R, Hodgkin P |title=Modeling T- and B-cell growth and differentiation |journal=Immunological Reviews |volume=216 |issue= |pages=119–29 |year=2007 |month=April |pmid=17367338 |doi=10.1111/j.1600-065X.2006.00498.x |url=http://www3.interscience.wiley.com/resolve/openurl?genre=article&sid=nlm:pubmed&issn=0105-2896&date=2007&volume=216&spage=119}}</ref><ref>{{cite journal |author=Hawkins ED, Hommel M, Turner ML, Battye FL, Markham JF, Hodgkin PD |title=Measuring lymphocyte proliferation, survival and differentiation using CFSE time-series data |journal=Nature Protocols |volume=2 |issue=9 |pages=2057–67 |year=2007 |pmid=17853861 |doi=10.1038/nprot.2007.297 }}</ref> Furthermore, the use of CFSE has extended beyond the immune system, with the dye being used to monitor the proliferation of many other cell types such as smooth muscle cells,<ref>{{cite journal |author=Sukkar MB, Stanley AJ, Blake AE, ''et al.'' |title='Proliferative' and 'synthetic' airway smooth muscle cells are overlapping populations |journal=Immunology and Cell Biology |volume=82 |issue=5 |pages=471–8 |year=2004 |month=October |pmid=15479432 |doi=10.1111/j.0818-9641.2004.01275.x }}</ref> fibroblasts,<ref>{{cite journal |author=Khil LY, Kim JY, Yoon JB, ''et al.'' |title=Insulin has a limited effect on the cell cycle progression in 3T3 L1 fibroblasts |journal=Molecules and Cells |volume=7 |issue=6 |pages=742–8 |year=1997 |month=December |pmid=9509415 |doi= |url=}}</ref> hematopoietic stem cells<ref>{{cite journal |author=Oostendorp RA, Audet J, Eaves CJ |title=High-resolution tracking of cell division suggests similar cell cycle kinetics of hematopoietic stem cells stimulated in vitro and in vivo |journal=Blood |volume=95 |issue=3 |pages=855–62 |year=2000 |month=February |pmid=10648396 |doi= |url=http://www.bloodjournal.org/cgi/pmidlookup?view=long&pmid=10648396}}</ref> and even bacteria!.<ref>{{cite journal |author=Ueckert JE, Nebe von-Caron G, Bos AP, ter Steeg PF |title=Flow cytometric analysis of Lactobacillus plantarum to monitor lag times, cell division and injury |journal=Letters in Applied Microbiology |volume=25 |issue=4 |pages=295–9 |year=1997 |month=October |pmid=9351280 |doi= |url=}}</ref> Another novel application of CFSE is its use for the ''in vitro'' and ''in vivo'' determination of cytotoxic lymphocytes.<ref>{{cite journal |author=Marzo AL, Kinnear BF, Lake RA, ''et al.'' |title=Tumor-specific CD4+ T cells have a major "post-licensing" role in CTL mediated anti-tumor immunity |journal=Journal of Immunology (Baltimore, Md. : 1950) |volume=165 |issue=11 |pages=6047–55 |year=2000 |month=December |pmid=11086036 |doi= |url=http://www.jimmunol.org/cgi/pmidlookup?view=long&pmid=11086036}}</ref><ref>{{cite journal |author=Jedema I, van der Werff NM, Barge RM, Willemze R, Falkenburg JH |title=New CFSE-based assay to determine susceptibility to lysis by cytotoxic T cells of leukemic precursor cells within a heterogeneous target cell population |journal=Blood |volume=103 |issue=7 |pages=2677–82 |year=2004 |month=April |pmid=14630824 |doi=10.1182/blood-2003-06-2070 |url=http://www.bloodjournal.org/cgi/pmidlookup?view=long&pmid=14630824}}</ref><ref>{{cite journal |author=Hermans IF, Silk JD, Yang J, ''et al.'' |title=The VITAL assay: a versatile fluorometric technique for assessing CTL- and NKT-mediated cytotoxicity against multiple targets in vitro and in vivo |journal=Journal of Immunological Methods |volume=285 |issue=1 |pages=25–40 |year=2004 |month=February |pmid=14871532 |doi=10.1016/j.jim.2003.10.017 |url=http://linkinghub.elsevier.com/retrieve/pii/S002217590300440X}}</ref><ref>{{cite journal |author=Stambas J, Doherty PC, Turner SJ |title=An in vivo cytotoxicity threshold for influenza A virus-specific effector and memory CD8(+) T cells |journal=Journal of Immunology (Baltimore, Md. : 1950) |volume=178 |issue=3 |pages=1285–92 |year=2007 |month=February |pmid=17237374 |doi= |url=http://www.jimmunol.org/cgi/pmidlookup?view=long&pmid=17237374}}</ref> Since the initial description of CFSE it has been used in thousands of immunological studies, an example of an early proliferation study in animals being described by Kurts et al.<ref>{{cite journal |vauthors=Kurts C, Kosaka H, Carbone FR, Miller JF, Heath WR |title=Class I-restricted cross-presentation of exogenous self-antigens leads to deletion of autoreactive CD8(+) T cells |journal=The Journal of Experimental Medicine |volume=186 |issue=2 |pages=239–45 |date=July 1997 |pmid=9221753 |pmc=2198972 |doi= 10.1084/jem.186.2.239}}</ref> However, perhaps the most important CFSE investigations have been those demonstrating that many of the effector functions of lymphocytes, such as ] production by ]s,<ref>{{cite journal |vauthors=Gett AV, Hodgkin PD |title=Cell division regulates the T cell cytokine repertoire, revealing a mechanism underlying immune class regulation |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=95 |issue=16 |pages=9488–93 |date=August 1998 |pmid=9689107 |pmc=21365 |doi= 10.1073/pnas.95.16.9488|bibcode=1998PNAS...95.9488G |doi-access=free }}</ref><ref>{{cite journal |vauthors=Bird JJ, Brown DR, Mullen AC |title=Helper T cell differentiation is controlled by the cell cycle |journal=Immunity |volume=9 |issue=2 |pages=229–37 |date=August 1998 |pmid=9729043 |doi= 10.1016/S1074-7613(00)80605-6|display-authors=etal|doi-access=free }}</ref> and antibody ] by ]s,<ref>{{cite journal |vauthors=Hodgkin PD, Lee JH, Lyons AB |title=B cell differentiation and isotype switching is related to division cycle number |journal=The Journal of Experimental Medicine |volume=184 |issue=1 |pages=277–81 |date=July 1996 |pmid=8691143 |pmc=2192686 |doi= 10.1084/jem.184.1.277}}</ref> are division dependent. Sophisticated mathematical models have also been developed to analyse CFSE data and probe various aspects of immune responses.<ref>{{cite journal |vauthors=Nordon RE, Nakamura M, Ramirez C, Odell R |s2cid=5696309 |title=Analysis of growth kinetics by division tracking |journal=Immunology and Cell Biology |volume=77 |issue=6 |pages=523–9 |date=December 1999 |pmid=10571673 |doi=10.1046/j.1440-1711.1999.00869.x }}</ref><ref>{{cite journal |vauthors=Gett AV, Hodgkin PD |title=A cellular calculus for signal integration by T cells |journal=Nature Immunology |volume=1 |issue=3 |pages=239–44 |date=September 2000 |pmid=10973282 |doi=10.1038/79782 |s2cid=8823323 }}</ref><ref>{{cite journal |vauthors=De Boer RJ, Ganusov VV, Milutinović D, Hodgkin PD, Perelson AS |title=Estimating lymphocyte division and death rates from CFSE data |journal=Bulletin of Mathematical Biology |volume=68 |issue=5 |pages=1011–31 |date=July 2006 |pmid=16832737 |doi=10.1007/s11538-006-9094-8 |hdl=1874/22578 |s2cid=6571349 |hdl-access=free }}</ref><ref>{{cite journal |vauthors=Callard R, Hodgkin P |title=Modeling T- and B-cell growth and differentiation |journal=Immunological Reviews |volume=216 |pages=119–29 |date=April 2007 |pmid=17367338 |doi=10.1111/j.1600-065X.2006.00498.x |s2cid=27565949 }}</ref><ref>{{cite journal |vauthors=Hawkins ED, Hommel M, Turner ML, Battye FL, Markham JF, Hodgkin PD |title=Measuring lymphocyte proliferation, survival and differentiation using CFSE time-series data |journal=Nature Protocols |volume=2 |issue=9 |pages=2057–67 |year=2007 |pmid=17853861 |doi=10.1038/nprot.2007.297 |s2cid=13550456 }}</ref> Furthermore, the use of CFSE has extended beyond the immune system, with the dye being used to monitor the proliferation of many other cell types such as ],<ref>{{cite journal |vauthors=Sukkar MB, Stanley AJ, Blake AE |title='Proliferative' and 'synthetic' airway smooth muscle cells are overlapping populations |journal=Immunology and Cell Biology |volume=82 |issue=5 |pages=471–8 |date=October 2004 |pmid=15479432 |doi=10.1111/j.0818-9641.2004.01275.x |s2cid=25686588 |display-authors=etal}}</ref> ]s,<ref>{{cite journal |vauthors=Khil LY, Kim JY, Yoon JB |title=Insulin has a limited effect on the cell cycle progression in 3T3 L1 fibroblasts |journal=Molecules and Cells |volume=7 |issue=6 |pages=742–8 |date=December 1997 |pmid=9509415 |display-authors=etal}}</ref> ]s<ref>{{cite journal |vauthors=Oostendorp RA, Audet J, Eaves CJ |title=High-resolution tracking of cell division suggests similar cell cycle kinetics of hematopoietic stem cells stimulated in vitro and in vivo |journal=Blood |volume=95 |issue=3 |pages=855–62 |date=February 2000 |pmid=10648396 |doi= 10.1182/blood.V95.3.855.003k41_855_862|url=http://www.bloodjournal.org/cgi/pmidlookup?view=long&pmid=10648396}}</ref> and even bacteria.<ref>{{cite journal |vauthors=Ueckert JE, Nebe von-Caron G, Bos AP, ter Steeg PF |title=Flow cytometric analysis of Lactobacillus plantarum to monitor lag times, cell division and injury |journal=Letters in Applied Microbiology |volume=25 |issue=4 |pages=295–9 |date=October 1997 |pmid=9351280 |doi= 10.1046/j.1472-765x.1997.00225.x|doi-access=free }}</ref> Another novel application of CFSE is its use for the ''in vitro'' and ''in vivo'' determination of ].<ref>{{cite journal |vauthors=Marzo AL, Kinnear BF, Lake RA |title=Tumor-specific CD4+ T cells have a major "post-licensing" role in CTL mediated anti-tumor immunity |journal=Journal of Immunology |volume=165 |issue=11 |pages=6047–55 |date=December 2000 |pmid=11086036 |doi= 10.4049/jimmunol.165.11.6047|display-authors=etal|doi-access=free }}</ref><ref>{{cite journal |vauthors=Jedema I, van der Werff NM, Barge RM, Willemze R, Falkenburg JH |s2cid=1984056 |title=New CFSE-based assay to determine susceptibility to lysis by cytotoxic T cells of leukemic precursor cells within a heterogeneous target cell population |journal=Blood |volume=103 |issue=7 |pages=2677–82 |date=April 2004 |pmid=14630824 |doi=10.1182/blood-2003-06-2070 |doi-access=free }}</ref><ref>{{cite journal |vauthors=Hermans IF, Silk JD, Yang J |title=The VITAL assay: a versatile fluorometric technique for assessing CTL- and NKT-mediated cytotoxicity against multiple targets in vitro and in vivo |journal=Journal of Immunological Methods |volume=285 |issue=1 |pages=25–40 |date=February 2004 |pmid=14871532 |doi=10.1016/j.jim.2003.10.017 |display-authors=etal}}</ref><ref>{{cite journal |vauthors=Stambas J, Doherty PC, Turner SJ |title=An in vivo cytotoxicity threshold for influenza A virus-specific effector and memory CD8(+) T cells |journal=Journal of Immunology |volume=178 |issue=3 |pages=1285–92 |date=February 2007 |pmid=17237374 |doi= 10.4049/jimmunol.178.3.1285|doi-access=free }}</ref>


Detailed protocols are now available that can be used to label lymphocytes (and other cell types) with a high degree of reliability and precision.<ref>{{cite journal |author=Lyons AB, Doherty KV |title=Flow cytometric analysis of cell division by dye dilution |journal=] |volume=Chapter 9 |issue= |pages=Unit 9.11 |year=2004 |month=February |pmid=18770808 |doi=10.1002/0471142956.cy0911s27 }}</ref><ref>{{cite journal |author=Quah BJ, Warren HS, Parish CR |title=Monitoring lymphocyte proliferation in vitro and in vivo with the intracellular fluorescent dye carboxyfluorescein diacetate succinimidyl ester |journal=Nature Protocols |volume=2 |issue=9 |pages=2049–56 |year=2007 |pmid=17853860 |doi=10.1038/nprot.2007.296 }}</ref><ref>{{cite journal |author=Parish CR, Glidden MH, Quah BJ, Warren HS |title=Use of the intracellular fluorescent dye CFSE to monitor lymphocyte migration and proliferation |journal=] |volume=Chapter 4 |issue= |pages=Unit4.9 |year=2009 |month=February |pmid=19235770 |doi=10.1002/0471142735.im0409s84 }}</ref> One of the most important parameters, however, is to ensure that the cell population being studied has not been too heavily labelled with CFSE as such cells, although remaining viable, proliferate sub-optimally. Detailed protocols are now available that can be used to label lymphocytes (and other cell types) with a high degree of reliability and precision.<ref>{{cite journal |vauthors=Lyons AB, Doherty KV |title=Flow cytometric analysis of cell division by dye dilution |journal=] |volume=Chapter 9 |pages=Unit 9.11 |date=February 2004 |pmid=18770808 |doi=10.1002/0471142956.cy0911s27 |isbn=0471142956 |s2cid=20204336 }}</ref><ref>{{cite journal |vauthors=Quah BJ, Warren HS, Parish CR |title=Monitoring lymphocyte proliferation in vitro and in vivo with the intracellular fluorescent dye carboxyfluorescein diacetate succinimidyl ester |journal=Nature Protocols |volume=2 |issue=9 |pages=2049–56 |year=2007 |pmid=17853860 |doi=10.1038/nprot.2007.296 |s2cid=1076080 }}</ref><ref>{{cite journal |vauthors=Parish CR, Glidden MH, Quah BJ, Warren HS |title=Use of the intracellular fluorescent dye CFSE to monitor lymphocyte migration and proliferation |journal=] |volume=Chapter 4 |pages=Unit4.9 |date=February 2009 |pmid=19235770 |doi=10.1002/0471142735.im0409s84 |isbn=978-0471142737 |s2cid=1953136 }}</ref> One of the most important parameters, however, is to ensure that the cell population being studied has not been too heavily labelled with CFSE, as such cells, although remaining viable, proliferate sub-optimally.


== References == == References ==
{{Reflist|2}} {{Reflist|2}}


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