Database of misidentified cell lines
R. Ian Freshney
Center for Oncology and Applied Pharmacology, Division of Cancer Sciences and Molecular Pathology, University of Glasgow,Glasgow, Scotland, United Kingdom

Letter to the Editor

Dear Sir,

Much of current cancer and cell biology research depends on
the use of cell lines cultured from normal and malignant tissue. However, ever since the time when continuous cell lines
were ﬁrst established, there has been a problem of the morevigorous lines contaminating and overgrowing more slowly
growing cultures. This has been compounded by confusion ofone cell line with another by mislabeling in routine culture orduring and after cryopreservation. The result is that some 15–20% of cell lines in current use may not be what they areclaimed to be. This has prompted a number of recent reports
in the literature1–7 and discussions at scientiﬁc meetings. Oneof the main conclusions is that there needs to be a way to alertscientists using established and frequently propagated cell linesthat there is a signiﬁcant risk that they may be using cell lines
which are not what they need them to be. This issue of International Journal of Cancer will address this problem andwants to increase the awareness of authors submitting theirwork for publication and of reviewers considering the merit ofthe work. Restrictions and conditions will be imposed regarding proof of authentication of cell lines used and advice given
on how to authenticate cell lines (see editorial and letter byW. Dirks). My purpose in this letter is to notify the scientiﬁccommunity of the existence and free availability of a list of celllines which are known or suspected to be falsely identiﬁed or
cross contaminated. This will allow scientists embarking on aproject or reviewers considering the work for publication, tohave access to a data source which will advise them on therespective cell line’s authenticity. This list is available fordownload from: http://www.hpacultures.org.uk/services/celllineidentityveriﬁcation/misidentiﬁedcelllines.jsp by following the link after my and Amanda Capes-Davis’s names. It
has been compiled from quality assurance carried out by anumber of cell banks (ATCC, CellBank Australia, sDSMZ,ECACC, JCRB, and RIKEN) and published on their websites,from an entry in Wikipedia, and from reports in the scientiﬁcliterature. It must be emphasized that while many of the celllines listed are clearly and incontrovertibly not what they aresupposed to be, original and authentic stocks of other lines
may yet exist. Where this is believed to be the case the line isincluded in the second table. This list will be published(Capes-Davis et al., ms in preparation).
I would request that anyone who uses this list and ﬁnds
that some misidentiﬁed cell lines have been omitted or thatsome cell lines reported as misidentiﬁed do have authentic
stocks available should contact me (i.freshney@ntlworld.com),and I will arrange to have the database updated.

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Int. J. Cancer: 126, 302–304 (2010) V

The recommended procedure for anyone contemplating
the use of cell lines is as follows:• Check that the cell line that you intend to use is notlisted in the above database.
that the cell line is obtained from a properly
authenticated source (and that may not be the originator), preferably from one of the recognized cell banks.
• Authenticate cell lines received from a nonauthenticatedsource on receipt (see letter of W. Dirks, this issue, andinstruction for authors of IJC).
• Repeat authentication at intervals of 3–6 months for celllines used for an extended study, before cryopreservation, and after thawing for further use.
• Ensure

It may not be possible to eliminate misidentiﬁcation
entirely, as new examples will continue to appear, but following these precautions should reduce the frequency and minimize the spread of the problem.
Yours sincerely,
R. Ian Freshney

References
1.

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4.
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7.

Stacey GN, Masters JRM, Hay RJ, Drexler HG, MacLeod RAF,Freshney RI. Cell contamination leads to inaccurate data: we musttake action now. Nature 2000;403:456.
Masters J. Re: false cell lines. Exp Cell Res 2002;272:216.
Drexler HG, Dirks WG, Matsuo Y, MacLeod RA. False leukemialymphoma cell lines: an update on over 500 cell lines. Leukemia.
2003;17:416–26.
Nardone RM. Curbing rampant cross-contamination andmisidentiﬁcation of cell lines. Biotechniques. 2008;45:221–7.
Lacroix M. Persistent use of ‘‘false’’ cell lines. Int J Cancer. 2008;122:
1–4.
MacLeod RAF, Dirks WG, Drexler HG. One falsehood leads easily
to another. Int J Cancer. 2008;122:2165–8.
Schweppe RE, Klopper JP, Korch C, Pugazhenthi U, Benezra M,
Knauf JA, Fagin JA, Marlow LA, Copland JA, Smallridge RC, Haugen
BR. Deoxyribonucleic acid proﬁling analysis of 40 human thyroid
cancer cell lines reveals cross-contamination resulting in cell line
redundancy and misidentiﬁcation. J Clin Endocrinol Metab. 2008;93:
4331–41.

DOI: 10.1002/ijc.24998
History: Received 3 May 2009; Accepted 12 May 2009; Online 23October 2009
Correspondence to: R. I. Freshney, 24 Greenwood Drive, Bearsden,
Glasgow G61 2HA, Scotland, United Kingdom,
E-mail: i.freshney@ntlworld.com

Letter to the Editor

303

Cell line cross-contamination initiative: an interactive reference
database of STR profiles covering common cancer cell lines
Wilhelm G. Dirks1, Roderick A. F. MacLeod1, Yukio Nakamura2, Arihiro Kohara3, Yvonne Reid4, Herbert Milch1,Hans G. Drexler1, Hiroshi Mizusawa3
1

DSMZ—Department of Human and Animal Cell Cultures, Braunschweig, GermanyRIKEN—Cell Engineering Division, Tsukuba, Japan
3
JCRB—Japanese Collection of Research Bioresources, Osaka, Japan
4
ATCC—American Type Culture Collections, Manassas, VA
2Recent reports1–4 demonstrate the growing perception in
the scientiﬁc community that cross contamination (CC) ofmammalian cell lines represents a major risk for generating
false scientiﬁc data. The level to which research has been
compromised by the use of contaminated or misidentiﬁed
cell lines has become a major concern for scientists, granting
agencies, and, increasingly, scientiﬁc journals. In 2007, agroup of cell biologists led by Roland M. Nardone petitioned
the United States Secretary of Health and Human Services todevelop an active program for cell line authentication.5 They
stressed that research and teaching tools in diverse ﬁelds ofscience and industry would be unimaginable without cell cultures. Despite the key importance of cell cultures, only little
consensus exists regarding the technical means by which cellline identity can be controlled and how to follow through theresults of any such testing.
The key problems of CC are known and chronic in nature: neglecting guidelines for quality control and disregarding adequate cell culture techniques are the main reasons
why cell lines have been misidentiﬁed or cross contaminated.
The incidence of CC in directly and indirectly provenanced
cell lines alike1,3 implies that the majority of false cell lines
are perpetrated in originators’ own laboratories, presumably
by failures during the establishment of new cell lines. A
plethora of reports unmasking bogus cancer cell lines, including members of the NCI-60 panel used to generate reference
baseline transcriptional drug responses has triggered calls for
remedial action.5,6 Nevertheless, standard authentication procedures for testing cell line identity have yet to be deﬁned.
Short tandem repeat (STR) microsatellite sequences arehighly polymorphic in human populations, and their stability
throughout the lifespan of individuals renders STR proﬁling(typing) ideal for forensic use. STR typing has served as areference technique for identity control of human cell lines at
Biological Resource Centers (BRCs) since the turn of the millennium.7 Ideally, authentication involves coincident STR
typing of paired donor and derived cell line samples. However, this ideal is met by a few recently established cell lines
only. Most widely used cell lines are decades old and their

C 2009 UICC
Int. J. Cancer: 126,302–304 (2010) V

identiﬁcation is largely retrospective and multidisciplinary,combining diverse criteria such as uniqueness and the congruence of STR proﬁles across independent samples with theconsistency of observed karyotypes with those reported by
the originators.
The DSMZ as well as the ATCC, JCRB, and RIKEN repositories have generated large databases of STR cell line proﬁles. By using the same microsatellite loci at these BRCs,
individual databases could be merged, thereby facilitatinginteractive searches. This work was piloted at the DSMZ togenerate an international reference STR proﬁle database forhuman cell lines. To render it user friendly, a simple searchengine for interrogating STR cell line proﬁles has now been
made available on the homepages of JCRB and DSMZ
(http://cellbank.nibio.go.jp/cellbank_e.html, http://www.dsmz.
de/STRanalysis). Registered users simply login at the searchsite on the DSMZ homepage and will be guided. Aided bysimple prompts, users can input their own cell line STR data
to retrieve best matches with authenticated cell lines listed on
the database.
Once the problem of false negatives due to discrepant representation of single STR alleles, e.g., by losses of heterozygosity and bottlenecking selection—has been tackled andunambiguous search results are produced, human cell lines
will need to be consistent with consensus STR reference datasets. STR proﬁles of all human cell lines distributed byDSMZ, JCRB, and RIKEN and one-third of the cell lines distributed by ATCC are now publicly accessible on interactive
databases where match criteria have been arbitrarily set to95%. Inevitably, reference proﬁles remain subject to revision
until all commonly held cell lines have been STR typed
across participating repositories. At present, about 2,342 suchcell lines have been STR typed and are represented as reference sets on the database.
The authors of this article are currently participating in
an international workgroup organized by the ATCC Standards Development Organization, (ATCC SDO) to develop astandardized methodology (protocols and procedures for STRanalysis) for authenticating human cell lines. An additional

Letter to the Editor

Dear Sir,

Letter to the Editor

304

goal of the workgroup is to establish a global database for
STR proﬁles of human cell lines. The development of theconsensus standard offers a new tool to the cell biology community that will foster reproducibility and comparability ofcell lines used in different laboratories. Armed with thesetools, online veriﬁcation of cell line identity should prove avital weapon to combat the havoc of cell line cross contamination which has dogged cancer research since inception.

Yours sincerely,Wilhelm G. Dirks
Roderick A. F. MacLeod
Yukio Nakamura
Arihiro Kohara
Yvonne Reid
Herbert Milch
Hans G. Drexler
Hiroshi Mizusawa

References
Drexler HG, Dirks WG, Matsuo Y, MacLeod RAF. False leukemialymphoma cell lines: an update on over 500 cell lines. Leukemia 2003;
17:416–26.

3.

4.

5.
6.

7.

Liscovitch M, Ravid D. A case study in misidentiﬁcation of cancer celllines: MCF-7/AdrR cells (re-designated NCI/ADR-RES) are derivedfrom OVCAR-8 human ovarian carcinoma cells. Cancer Lett 2007;
245:350–52.
MacLeod RAF, Dirks WG, Kaufmann M, Matsuo Y, Milch H, Drexler
HG. Widespread intra-species cross-contamination of human tumor cellline arising at source. Int J Cancer 1999;83:555–63.
Schweppe RE, Klopper JP, Korch C, Pugazhenthi U, Benezra M,
Knauf JA, Fagin JA, Marlow LA, Copland JA, Smallridge RC, Haugen
BR. Deoxyribonucleic acid proﬁling analysis of 40 human thyroid
cancer cell lines reveals cross-contamination resulting in cell line
redundancy and misidentiﬁcation. J Clin Endocrinol Metab 2008;93:4331–41
Nardone RM. Eradication of cross-contaminated cell lines: a call foraction. Cell Biol Toxicol 2007;23:367–72.
Stacey, GN, Masters, JRM, Hay, RJ, Drexler, HG, MacLeod, RAF,Freshney, RI. Cell contamination leads to inaccurate data: we musttake action now. Nature 2000;403:456.
Masters JR, Thompson JA, Daly-Burns B, et al. Short tandem repeat
proﬁling provides an international reference standard for human celllines. Proc Natl Acad Sci 2001;98:8012–7.

DOI: 10.10002/ijc.24999History: Received 24 Jun 2009; Accepted 16 Oct 2009; Online 26
October 2009
Correspondence to: W. G. Dirks, DSMZ—Department of Humanand Animal Cell Culture, Braunschweig, Germany,
E-mail: wdi@dsmz.deLetter to the Editor

1.

2.

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Int. J. Cancer: 126, 302–304 (2010) V