-Detailed Information [JCRB0601]-

[問題が該当する細胞株]
JCRB0601 BALB/3T3 A31-1-1
IFO50070 Balb/3T3-A31-1-1
IFO50299 Balb/c 3T3 A31-1-13
IFO50298 Balb/c 3T3 A31-I-1
JCRB0149 Bhas 42
JCRB1355 1-1ras1000
JCRB1356 A31-1-1
JCRB1357 1-1 src
	
これらの細胞は、マウス系統解析から、Swissマウスに由来する細胞であり、Balb/c 3T3 clone A31に由来する細胞ではないことが判明しました。

Balb/c 3T3 A31-1-1, Balb/c 3T3 A31-1-13細胞は、親株をBalb/c 3T3 clone A31細胞とするサブクローンとして、角永先生の研究室（当時National Cancer Intitute）で樹立されました。これらのサブクローンは、親株のA31よりも化学発癌物質によるトランスフォーメーションに適するものとされ、世界的に広く利用されています。JCRB細胞バンクにも複数のルートから寄託されました。なお、IFO50070番で登録された株は、角永先生から直接バンクに寄託されたものです。
その後、さらに他の研究室によりBalb/c 3T3 A31-1-1を親株として遺伝子導入された細胞株、Bhas 42, 1-1ras1000, 1-1 srcが樹立され、JCRB細胞バンクに寄託されました。

しかし、JCRB細胞バンクが細胞の由来を確かめる目的で、これらの細胞株のマウス系統解析を行ったところ、角永先生の研究室で樹立されたこれらのサブクローン、および、A31-1-1に派生する細胞株はすべてSwissマウス由来であることがわかりました。

一方、元々の親株とされるBalb/c 3T3 clone A31細胞は、正しくBalb/cマウス由来であることが確認されました。

角永先生はすでにかなり前に逝去されており、どのような状況でこのような事態が生じたのかは確認ができませんが、Balb/c 3T3 A31-1-1 A31-1-13は、何らかの混同、あるいは誤認によりSwissマウスの線維芽細胞をクローンとして拾って樹立した株ということになります。

これらの細胞株は、歴史的にも広く用いられてきた経緯、あるいは試験法に記載されてきた経緯等があり、その有用性は失われておりません。このため、JCRB細胞バンクでは、これらの細胞の登録を抹消せず、分譲を継続しています。しかし、上記細胞の使用に関しては、Balb/c 3T3 clone A31細胞とは系統が異なる細胞であることをご了解ください。

例えば、もし、枝番がついていないA31細胞としての性質を求めるのであれば（たとえばOECDの光毒性試験など)、JCRB9005で登録されているBALB/3T3 clone A31のご利用をご検討ください。

一方、角永らにより樹立された、トランスフォームしやすいとされる細胞のご利用をお考えの場合、または、論文の追試や、試験法などで細胞名が明記されている場合には、マウス系統の問題はあるものの、その使用は妥当となります。

つきましては、細胞のご研究への適合性についてご留意くださいますようお願い申し上げます。

参考文献：In Vitro Cell Dev Biol Anim. http://link.springer.com/article/10.1007%2Fs11626-016-0104-3


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[The cell lines in problem]

JCRB0601 BALB/3T3 A31-1-1
IFO50070 Balb/3T3-A31-1-1
IFO50299 Balb/c 3T3 A31-1-13
IFO50298 Balb/c 3T3 A31-I-1
JCRB0149 Bhas 42
JCRB1355 1-1ras1000
JCRB1356 A31-1-1
JCRB1357 1-1 src

These cell lines have been proved to be derived from Swiss mouse cells, but not from the derivative of Balb/c 3T3 clone A31.

Balb/3T3 A31-1-1 and A31-1-13 were reported to be isolated from Balb/3T3 clone A31 by Dr. Kakunaga at National Cancer Institute in 1980 (Science 209: 505-507, 1980). These cell lines were reported to be useful for the study of chemically induced transformation of cells, and were widely used in the world. These cell lines were deposited at JCRB Cell Bank from different deposition routes. For example, IFO50070 strain was the one that was directly deposited by Dr. Kakunaga.

Thereafter, several laboratories developed genetically modified cell lines from Balb/c 3T3 A31-1-1. These are Bhas 42, 1-1ras1000, 1-1 src, and were also deposited at JCRB Cell Bank.

JCRB Cell Bank took place an analysis of mouse strain to check the origin of these cell lines. During this analysis, we found that A31-1-1, A31-1-13 and all these derivatives are NOT derived from Balb/c mouse, but from Swiss mouse.

Wheras, the parental cell line, Balb/c 3T3 clone A31 was confirmed as Balb/c mouse origin.

Dr. Kakunaga passed away in 1988, and unfortunately we can not trace the situation and cause of problem on this matter. However, it is suspected that the Balb/3T3 A31-1-1 and A31-1-13 subclones used in his laboratory was mistakenly establihed (cloned) from Swiss mouse-derived fibroblasts.

These cell lines have been historically used in worldwide, or have been described in test guidelines, and therefore the still valuable for research and testing. For this reason, JCRB does not exclude these cell lines from catalog and continues distribution of them. However, please keep in mind that these cell lines are not Balb/3T3 A31-derived cells.

For example, If your research needs the characteristics of Balb/c 3T3 clone A31 (such as OECE test guideline "In vitro 3T3 NRU phototoxicity test"), please consider to use JCRB9005 BALB/3T3 clone A31.

Wheras if you will use the cells established by Kakunaga for transformation studies, or if the name of cell lines are clearly described in the paper or test guidelines, these cell lines are still available for use regardless to the mouse strain problem.

Sincerely,

JCRB Cell Bank

References: In Vitro Cell Dev Biol Anim. Http://link.springer.com/article/10.1007%2 Fs 11626-016-0104-3
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LOT Information

JCRB No.	JCRB0601	Cell Name	BALB/3T3 A31-1-1
Profile	fibroblast, contact inhibited, (Strain problem was found. Refer Important Notice at cell detail page.)	Other Name	A31-1-1
Animal	mouse	Strain	Swiss
Genus	Mus	Species	musclus
Sex		Age	embryo
Identity		Tissue for Primary Cancer	embryo
Case history	normal	Metastasis
Tissue Metastasized		Genetics	Origin of this cell line, Balb/c 3T3 A31-1, was found not to be derived from BALB 3T3 A31.
Life Span	infinite	Crisis PDL
Morphology	fibroblast-like	Character	contact inhibited mouse cell line
Classify	normal	Established by	Kakunaga,T.
Registered by	Kuroki,T.	Regulation for Distribution
Comment	The FCS requires careful selection to keep quality.	Year
Medium	Eagle's minimal essential medium with 10% fetal calf serum	Methods for Passages	0.25% trypsin & 0.02% EDTA. Practically, 1/100 dilution once a week. Avoid being confluent.
Cell Number on Passage	3x10^5 cells/60 mm dish	Race
CO2 Conc.	5 %	Tissue Sampling	embryo
Tissue Type

Detection of virus genome fragment by Real-time PCR
Detected DNA Virus	not tested	Detected RNA Virus	not tested

Link
Cellosaurus(ExPASy) Cellosaurus (ExPASy) is developed by Amos Bairoch of the CALIPHO group at the SIB - Swiss Institute of Bioinformatics as part of the neXtProt project. Cellosaurus is a knowledge database of cell lines with various information summarized.	CVCL_J646

Reference
Pubmed id:7394516	Cell variants showing differential susceptibility to ultraviolet light--induced transformation. Kakunaga T,Crow JD Science. 1980 Jul 25;209(4455):505-7
Pubmed id:4311790	Murine sarcoma and leukemia viruses: assay using clonal lines of contact-inhibited mouse cells. Jainchill JL,Aaronson SA,Todaro GJ J Virol. 1969 Nov;4(5):549-53
Pubmed id:4301006	Development of 3T3-like lines from Balb-c mouse embryo cultures: transformation susceptibility to SV40. Aaronson SA,Todaro GJ J Cell Physiol. 1968 Oct;72(2):141-8
Pubmed id:13985244	Quantitative studies of the growth of mouse embryo cells in culture and their development into established lines. TODARO GJ,GREEN H J Cell Biol. 1963 May;17():299-313

09202011
05142008
043096
021486
950821

Cell No.	JCRB0601	Cell Name
LOT No.	09202011	Lot Specification	distribution
Medium	Eagle's minimum essential medium with 10% fetal bovine serum (FBS: GIBCO Cat. # 10099)	Temperature	37 C
Cell Density at Seeding	approx. 1 x 10^4 cells/mL	Methods for Passages	Cells were harvested after treatment with 0.25% trypsin and 0.02% EDTA
Doubling Time	approx. 1 day	Cell Number in Vial (cells/1ml)	1.5 x 10^6
Viability at cell freezing (%)	97.3	Antibiotics Used	free
Passage Number	Unknown (6 at benk)	PDL
Sterility: MYCOPLASMA	-	Sterility: BACTERIA	-
Sterility: FUNGI	-	Isozyme Analysis	Confirmed as mouse by NP, G6PD, MD.
Chromosome Mode		Chromosome Information
Surface Antigen		DNA Profile (STR)
Adhesion	Yes	Exoteric Gene
Medium for Freezing	10% DMSO, 20% FBS - EMEM	CO2 Conc.	5%
Viability immediately after thawing (%)		Additional information

Images

Cell No.	JCRB0601	Cell Name	Balb/3T3-A31-1-1
LOT No.	05142008	Lot Specification	distribution
Medium	Eagle's minimum essential medium with 10% fetal bovine serum (FBS Lot; GIBCO Cat. No. 10099, Lot 484375) // The number on the label is misprinted: the correct number is JCRB0601.	Temperature	37 C
Cell Density at Seeding	1 x 10^4 cells/ml	Methods for Passages	0.25% trypsin and 0.02% EDTA. Passage twice a week. Do not allow the culture to be full confluency.
Doubling Time	18 hrs	Cell Number in Vial (cells/1ml)	3.5 x 10^6
Viability at cell freezing (%)	98.8	Antibiotics Used	free
Passage Number	Unknown (6 at bank)	PDL
Sterility: MYCOPLASMA	-	Sterility: BACTERIA	-
Sterility: FUNGI	-	Isozyme Analysis	Confirmed as mouse by NP, G6PD, MD.
Chromosome Mode		Chromosome Information
Surface Antigen		DNA Profile (STR)
Adhesion	Yes	Exoteric Gene
Medium for Freezing	10% DMSO, 20% FBS - EMEM	CO2 Conc.	5%
Viability immediately after thawing (%)		Additional information

Images

Cell No.	JCRB0601	Cell Name	BALB/3T3 A31-1-1
LOT No.	043096	Lot Specification	distribution
Medium		Temperature
Cell Density at Seeding		Methods for Passages
Doubling Time		Cell Number in Vial (cells/1ml)
Viability at cell freezing (%)		Antibiotics Used
Passage Number	P6*	PDL
Sterility: MYCOPLASMA	-	Sterility: BACTERIA	-
Sterility: FUNGI	-	Isozyme Analysis	Confirmed as a mouse by LDH, NP and G6PD.
Chromosome Mode		Chromosome Information
Surface Antigen		DNA Profile (STR)
Adhesion		Exoteric Gene
Medium for Freezing		CO2 Conc.
Viability immediately after thawing (%)		Additional information

Images

Cell No.	JCRB0601	Cell Name	BALB/3T3 A31-1-1
LOT No.	021486	Lot Specification	distribution
Medium	MEM + FCS 10%	Temperature	37 C
Cell Density at Seeding	3x10^5 cells/60 mm dish	Methods for Passages	3T3 (3x10^5/60mm D every 3 days). Practically, 1:100 dilution, once a week. Never use for TF if pass.
Doubling Time	NT	Cell Number in Vial (cells/1ml)	1.3x10^6
Viability at cell freezing (%)	94%	Antibiotics Used	free
Passage Number	P7	PDL
Sterility: MYCOPLASMA	-	Sterility: BACTERIA	-
Sterility: FUNGI	-	Isozyme Analysis	AST,G6PD,LD,MD,MPI,NP,PEPB
Chromosome Mode		Chromosome Information	NT
Surface Antigen		DNA Profile (STR)
Adhesion		Exoteric Gene
Medium for Freezing		CO2 Conc.
Viability immediately after thawing (%)		Additional information

Cell No.	JCRB0601	Cell Name
LOT No.	950821	Lot Specification	distribution
Medium	MEM + FCS 10%	Temperature	37
Cell Density at Seeding	3x10^5/60 mm D	Methods for Passages	3T3 (3x10^5/60mm D every 3 days). Practically, 1:100 dilution, once a week.
Doubling Time		Cell Number in Vial (cells/1ml)
Viability at cell freezing (%)		Antibiotics Used	kanamycin 60 ug/ml
Passage Number		PDL
Sterility: MYCOPLASMA		Sterility: BACTERIA
Sterility: FUNGI		Isozyme Analysis
Chromosome Mode		Chromosome Information
Surface Antigen		DNA Profile (STR)
Adhesion		Exoteric Gene
Medium for Freezing		CO2 Conc.
Viability immediately after thawing (%)		Additional information

JCRB Cell Bank

JCRB0601 BALB/3T3 A31-1-1

Cell information

LOT Information