A-Cube®

Academicmaterial

ProteoBridge Co., Ltd.
Clinical Laboratory Center

2022.12 RevisedInstructions for Research and Examination
Examination Items

A-CubeSystemic Sclerosis
Related Antibodies
Detection Kit（SSc）
33 Antigens

A-CubeDermatomyositis/
Polymyositis Related
Antibodies Detection
Kit（DM/PM）
47 Antigens

Detection antibody（antigen）
CENP-A（CENPA）,CENP-B（CENPB）,CENP-C（CENPC）,
Scl-70/TopoⅠ（TOP1）,RNAPⅢ/RPC155（POLR3A）,
RNAPⅢ/RPC62（POLR3C）,RNAPⅠ（POLR1A）,RNAPⅡ（POLR2A）,
Th/To/7-2RNP（POP1,RPP25）,U3-RNP/Fibrillarin（ FBL）,hUBF/NOR90（UBTF）,U11/U12-RNP（RNPC3）,SSSCA1（SSSCA1）,eIF2B（EIF2B2）,AMA-M2/Mitochondria M2（DLAT,DLST,DBT,PDHX）,
p80-coilin（ COIL）

U1-RNP̲70（SNRNP70）,U1-RNP̲A（SNRPA）,U1-RNP̲C（SNRPC）,
U2-RNP（SNRPB2）,Ku（ XRCC5,XRCC6）,
PM-Scl100（EXOSC10）,PM-Scl75（EXOSC9）,RuvBL1/2（RUVBL1&2）,
Ki（PSME3）,SS-A/Ro52（TRIM21）,SS-A/Ro60（TROVE2）,SS-B（SSB）
Jo-1（HARS）,PL-7（TARS）,PL-12（AARS）,EJ（ GARS）,KS（NARS）,
OJ（ IARS,EPRS,LARS,MARS,QARS,KARS,RARS,DARS,AIMP1,2,3）,Zo（ FARSA,FARSB）,Ha（ YARS）,
SRP（SRP54,14,19,68,72）,Mi-2（CHD3,CHD4）,
TIF1-γ/p155（TRIM33）,TIF1-α/p140（TRIM24）,TIF1-β（TRIM28）,
MJ/NXP-2（MORC3）,SAE（SAE1,UBA2）,SMN（SMN1）,cN1A（NT5C1A）

U1-RNP̲70（SNRNP70）,U1-RNP̲A（SNRPA）,U1-RNP̲C（SNRPC）,
U2-RNP（SNRPB2）,Ku（ XRCC5,XRCC6）,
PM-Scl100（EXOSC10）,PM-Scl75（EXOSC9）,RuvBL1/2（RUVBL1&2）,
Ki（PSME3）,SS-A/Ro52（TRIM21）,SS-A/Ro60（TROVE2）,SS-B（SSB）

A-CubeMultiplex Test Kit（DM/PM + SSc Related

SSc-only 20 antigens + DM/PM only 34 antigens + Common
13 antigens total 67 antigensAntibodies）67 Antigens※Italic:detectable antibodies（antigens）shared between SSc and DM/PM

This is an indirect ﬂuorescent antibody test that uses nondried antigenic proteins and can detect multiple
autoantibodies individually and comprehensively at a time from small amounts of serum to conventional products.
This test allows detection of autoantibodies associated with systemic scleroderma and dermatomyositis/polymyositis,
and the results of the test report the quantitative value（index value) of antibodies to each antigen.
※This test chip was developed and named A-Cube（Autoantibody Array Assay）under the supervision ofDr. Yoshizaki and Professor Sato of Dermatology, Graduate School of Medicine, University of Tokyo.
※This test technology was developed by the National Institute of Industrial Technology, and the test results
will be provided by ProteoBridge Co., Ltd., an AIST venture.
Serum
Fluorescent labeling
Antihuman IgG antibody

Autoantibodies
Antigens1

List of detected antibodies and antigens
Detection antibody

AntigenDetection antibody

Antigen

CENP-A

CENPA

Jo-1

HARS

CENP-B

CENPB

PL-7

TARS

CENP-C

CENPC

PL-12

AARS

Scl-70（TopoⅠ）

TOP1

EJ

GARS

RNAPⅢ
（RPC155）

POLR3A

KS

NARS

RNAPⅢ
（RPC62）

POLR3C

IARS

RNAPⅠ

POLR1A

EPRS

RNAPⅡ

POLR2ALARS

POP1MARS

RPP25

QARS

Th/To（ 7-2RNP）
U3-RNP（Fibrillarin）

FBL

hUBF（NOR90）

UBTF

RARS

U11/U12-RNP

RNPC3DARSSSSCA1SSSCA1AIMP1eIF2B

EIF2B2

AIMP2DLAT

AIMP3

AMA-M2（Mitochondria M2）

DLST
DBT
PDHX

OJ

Zo
Ha

KARS

FARSA
FARSB
YARS

p80-coilin

COIL

SRP54

U1-RNP̲70SNRNP70

SRP14U1-RNP̲A

SNRPA

U1-RNP̲C

SNRPC

SRP68

U2-RNPSNRPB2

SRP72

XRCC5Ku

XRCC6SRP

Mi-2

SRP19

CHD3CHD4PM-Scl100

EXOSC10TIF1-γ（p155）

TRIM33PM-Scl75

EXOSC9TIF1-α（p140）

TRIM24

RuvBL1/2

RUVBL1&2

TIF1-βTRIM28Ki

PSME3

MJ（ NXP-2）

MORC3

SS-A/Ro52

TRIM21

SS-A/Ro60

TROVE2

SS-B

SSB

SAE

SAE1
UBA2

SMN

SMN1

cN1A

NT5C1A＋Systemic Sclerosis Related Antibodies Detection Kit（SSc）33 Antigens＋Dermatomyositis/Polymyositis Related Antibodies Detection Kit（DM/PM）47 Antigens＋＋Multiplex Test Kit（DM/PM + SSc Related Antibodies）67 Antigens
2

Systemic Sclerosis Related Antibodies Detection Kit（SSc）
More than 90% of patients with systemic scleroderma are positive for antinuclear antibodies. Amongthese antinuclear antibodies, anti-centromere antibodies, anti-topoisomerase I antibodies, and anti-RNA
polymerase III antibodies are covered as speciﬁc and representative of systemic scleroderma and areroutinely used in scleroderma diagnosis and treatment decision making.
In this kit, systemic scleroderma-speciﬁc antibodies can be detected in RNAPI, RNAPII, Th/To, U3-RNP（Fibrillarin）, hUBF（NOR90）, U11/U12-RNP, and eIF2B that cannot be measured by insurance-listed tests
other than those listed above. These autoantibodies have also been reported to be closely related to the clinicalpresentation.
Systemic scleroderma-related antibodies include anti-U1RNP antibodies（70, A, and C）, anti-U2RNP antibodies,anti-Ku antibodies, anti-PM-Scl antibodies（100, 75）, anti-RuvBL1/2 antibodies, and anti-Ki antibodies detectedby overlap with myositis.
,They can also detect anti-SSSCA1 （P27） antibodies, which are detected in scleroderma and Sjögren ssyndrome, and anti-AMA-M2（mitochondrial）antibodies（four antigens）and anti-p80-coilin antibodies, which aredetected in primary biliary cirrhosis, which is highly complicated with scleroderma.
As such, this test provides important information for typing, selecting treatment, and predicting prognosis because
speciﬁc and related autoantibodies for systemic scleroderma can be comprehensively measured at one time.

Performance correlation data
Percent agreement with samples determined by immunoprecipitation（mean of three autoantibodies each）.
●Anti-Th/To Ab●Anti-RNA-polymerase I/III/II Ab●Anti-U3-RNP Ab●Anti-Ku Ab●Anti-NOR90 Ab●Anti-SS-A Ab●Anti-PM-Scl Ab●Anti-SS-B Ab

Concordance rate̲100%（25/25）●Anti-topoisomerase I（Scl-70）Ab

Concordance rate with insurance-listed kits
96 samples
Correlation coeﬃcient r=0.942

1000
800

y=3.7336x−12.075
R2=0.8875

600
400
200
0050100▶Anti-Scl-70 Ab
This product（index value）

This product（index value）▶Anti-CENP-B Ab

150200

Products covered by insurance（index value）

300

y=0.8194x−3.7732
R2=0.821

250200150100500050100

A-Cube（This Product）

±3＋

200

1±01

1−0073200

250

ELISA（insured products）−＋Concordance rate̲98%

150

Products covered by insurance（index value）

ELISA（insured products）＋

98 samples
Correlation coeﬃcient r=0.906

A-Cube（This Product）

±−＋

29

1±000−1167

Concordance rate̲98%0Reference

Systemic Sclerosis Related Antibodies Detection Kit（SSc）
Antibody

Contents of the report

CENP-A

30% positive rate in scleroderma, localized scleroderma,skin ulcer, calciﬁcation, reﬂux esophagitis,
pulmonary hypertension correlated with good prognosis

CENP-B
CENP-C

Reference
Japanese Journal of Clinical Immunology,36（3）
139-147（2013）

40% positive rate in scleroderma,

Japanese Journal of Clinical Immunology,（3）139-147（2013）（pulmonary ﬁbrosis）, correlated with renal crisis,poor prognosis 36

Scl-70（TopoⅠ） diﬀuse type of skin sclerosis,interstitial pneumonia
RNAPⅢ（RPC１５５）
RNAPⅢ（RPC６２） 20% positive rate in scleroderma,
RNAPⅠ

diﬀuse type of skin sclerosis,
correlated with renal crisis, good prognosis

Japanese Journal of Clinical Immunology,36（3）
139-147（2013）

RNAPⅡ
2−5% positivity in scleroderma, localized scleroderma,

Th/To（7-2RNP） interstitial pneumonia（pulmonary ﬁbrosis）,
and pulmonary arterial hypertension

U3-RNP（Fibrillarin）

4−10% positive rate in scleroderma, diﬀuse type of skinsclerosis and peripheral circulatory disturbances, interstitial
pneumonia (pulmonary ﬁbrosis), pulmonary arterial
hypertension, correlate with lower gastrointestinal lesions
10% positive rate in rheumatic diseases, 3.3% of them have

（ＮＯＲ９０） scleroderma, localized skin sclerosis, correlate withｈＵＢＦ
non-serious visceral lesions

Japanese Journal of Clinical Immunology,36（3）
139-147（2013）
Japanese Journal of Clinical Immunology,36（3）
139-147（2013）
Arthritis Rheum.1996 Aug;39（8）：1313-8.
Japanese Journal of Clinical Immunology,36（3）
139-147（2013）

3.2% positivity in scleroderma, correlated withinterstitial pneumonia（pulmonary ﬁbrosis）

Arthritis Rheum. 2009 July 15;61（7）：958‒965.

SSSCA11.7% positivity for all autoimmune diseases,,
all positive for scleroderma or Sjögren s syndrome,and a subset of anti-CENP antibodies

Clin Exp Immunol. 1998 Feb;111（2）：372-6.

eIF2B

A positive rate of 1−2% in scleroderma,diﬀuse scleroderma
and interstitial lung disease were frequent,overlapping
myositis or rheumatoid arthritis, anticytoplasmic antibodies

Arthritis Rheum. 2016 Nov;68（11）:2778-2783.

U11/U12-RNP

AMA-M2
With a positive rate of 14.8% in scleroderma, an indicator of（Mitochondria M2） concomitant primary biliary cirrhosis（PBC）
p80-coilin
U1-RNP̲70U1-RNP̲ AU1-RNP̲C

Positive rates of primary biliary cirrhosis（PBC）
̲3.3% and,
Sjögren s syndrome̲3.7% across autoimmune diseases.

Kanzo, Vol.48 No.5 210-218（2007）J Exp Med. 1991 Jun 1;173（6）：1407-19.

2−14% positive rate for MCTD, DM/PM or SLE overlap syndrome,
Japanese Journal of Clinical Immunology,
sausage-like ﬁnger inﬂation, localized skin sclerosis,
36（3）139-147（2013）,Raynaud s phenomenon, arthritis, reﬂux esophagitis is common.

U2-RNP

Polymyositis-scleroderma overlap syndrome with lessthan 5% positivity, muscle weakness, and mild scleroderma

Clinical Rheumatology and Related Research,
25：149〜158,2013

Ku

Polymyositis-scleroderma overlap syndrome with 2−30% positivity,
muscle weakness, minimal skin sclerosis, and good vital prognosis

Clinical Rheumatology and Related Research,
25：149〜158,2013

2% positive rate in scleroderma, localized skin sclerosis,
less frequent serious visceral complications

Japanese Journal of Clinical Immunology,36（3）
139-147（2013）

RuvBL1/2

A positive rate of 1−2% in scleroderma, Overlap ofmyositis and diﬀuse skin thickening occur frequently,
higher incidence in male and elderly patients

Arthritis Care Res（Hoboken）.
2014 Apr;66（4）:575-84.

Ki

10% positivity rate in overlap syndromes according
to each collagen disease

Laboratory Medicine Encyclopedia & Dictionary

10−30% positivity in scleroderma

Laboratory Medicine Encyclopedia & Dictionary

Positive rate of 5% or less in scleroderma

Laboratory Medicine Encyclopedia & Dictionary

PM-Scl100
PM-Scl75

SS-A/Ro52
SS-A/Ro60
SS-B4Dermatomyositis/Polymyositis Related Antibodies
Detection Kit（DM/PM）
In dermatomyositis/polymyositis（hereafter myositis）, autoantibodies against various cellular components aredetected at a high rate. Among them, myositis-speciﬁc autoantibodies are clinically useful, such as diagnosing,classifying disease types, estimating prognosis, and determining treatment. Moreover, many of these
corresponding antigens have been elucidated as enzymes and regulators involved in important vital phenomena
such as gene transcription/translation and DNA repair, which provide important information for considering themechanism of autoantibody production and the pathogenesis of myositis.
Eight anti-ARS antibodies have been reported that are speciﬁc to myositis and autoantibodies to aminoacyl tRNA,
synthetase, with common clinical manifestations such as interstitial pneumonitis, fever, arthritis, Raynaud sdisease, and mechanics hands. In this kit, all eight types of Jo-1, PL-7, PL-12, EJ, KS, OJ, Zo, Ha can bedetected. Especially for anti-OJ antibodies, other than the major antigen, IARS, 10 other antigens that arecomplexed in vivo can also be detected. To date, there are no assay reagents that can detect antibodies against
these at one time, and this reagent enables detection equivalent to immunoprecipitation.
Other myositis-speciﬁc antibodies include anti-SRP antibodies, which are correlated with immune-mediatednecrotizing myopathy; anti-Mi-2 antibodies, which have high serum CK levels and often have a typical clinicalform of dermatomyositis; and anti-TIF1 antibodies（γ, α, and β）and anti-MJ antibodies（NXP-2）, which arecorrelated with malignancy in adults.
In addition, antibodies against SMNs that interact with snRNP, which have been reported as novel
myositis-speciﬁc antibodies, can also be detected. Myositis-related antibodies include anti-U1RNP antibodies（70, A, and C）, anti-U2RNP antibodies, anti-Ku antibodies, anti-PM-Scl antibodies（100,75）, anti-RuvBL1/2antibodies, and anti-Ki antibodies, which are detected by overlap with systemic scleroderma.

Performance correlation data
Percent agreement with samples determined by immunoprecipitation（mean of three autoantibodies each）.
●Anti-aminoacyl tRNA synthase（ARSs）Abs ●Anti-TIF1-β-Abs（Jo-1, PL-7, PL-12, EJ, KS, OJ）●Anti-NXP-2 Ab

Concordance rate̲98%（41/42）●Anti-SRP Ab●Anti-Mi-2 Ab●Anti-SAE Abs●Anti-SS-A Ab●Anti-TIF1-alpha/gamma Abs●Anti-SS-B Ab*Discordance : 1 specimen tested positive
for anti-TIF1β antibodies by immunoprecipitation→ negative by this productConcordance rate with insurance-listed kits

This product（index value）▶Anti-ARS Ab

26 samples
Correlation coeﬃcient r=0.933

200＋R2=0.8711

150A-Cube100−＋140±00−

111Concordance rate̲96%50*Since the comparator uses a mixture antigen against Jo-1,PL-7, PL-12, EJ, KS, product also produces and compares
mixture antigens for this comparison.

0050

100150

Products covered by insurance（index value）
5ELISA（Insurance listed items）

y=0.7725x−3.2786

200Reference

Dermatomyositis/Polymyositis Related Antibodies
Detection Kit（DM/PM）
Antibody

Contents of the report

Reference

ARS

15−20% positivity for myositis, interstitial pneumonia,,
polyarthritis, Raynaud s phenomenon, fever, mechanics
hands are more frequentClinical Rheumatology and Related Research,
25：149〜158,2013

Jo-1

15−20% positivity for myositis, polymyositis > dermatomyositis

Clinical Rheumatology and Related Research,
25：149〜158,2013

PL-7

Less than 5% positivity in myositis

Clinical Rheumatology and Related Research,
25：149〜158,2013

PL-12

Positive rate of less than 5% in myositis,
interstitial pneumonia > myositis

Clinical Rheumatology and Related Research,
25：149〜158,2013
Clinical Rheumatology and Related Research,
25：149〜158,2013
Clinical Rheumatology and Related Research,
25：149〜158,2013

EJ

5−10% positivity for myositis, dermatomyositis > polymyositis

KSPositive rate of less than 5% in myositis,interstitial pneumonia > myositis

OJ

Less than 5% positivity in myositis

Zo

Less than 1% positivity in myositis

Ha

Less than 1% positivity in myositis

SRPPolymyositis with a positive rate of 5%, severe/refractory/
recurrent/necrotizing myopathy, malignancy, and othercollagen diseases are less common

Clinical Rheumatology and Related Research,
25：149〜158,2013

Mi-2

5−10% positivity in dermatomyositis, low frequency ofinterstitial pneumonia and arthritis, and good steroid response

Clinical Rheumatology and Related Research,
25：149〜158,2013

20% positivity in dermatomyositis, signiﬁcantly more

Clinical Rheumatology and Related Research,
25：149〜158,2013

TIF1-γ（p155） frequently complicated with malignancy

10% positive rate in dermatomyositis, signiﬁcantly

TIF1-α（p140） more frequently complicated with malignancy
TIF1-βClinical Rheumatology and Related Research,
25：149〜158,2013
Clinical Rheumatology and Related Research,
25：149〜158,2013
Clinical Rheumatology and Related Research,
25：149〜158,2013Arthritis Rheum.2012 Feb;64（2）:513-22.

1.5% positivity in dermatomyositis

Arthritis Rheum.2012 Feb;64（2）:513-22.

Less than 5% positivity for dermatomyositis,and 27−32% positivity for children

Clinical Rheumatology and Related Research,
25：149〜158,2013

SAE

Positive rate of less than 1% in dermatomyositis, dysphagia

Clinical Rheumatology and Related Research,
25：149〜158,2013

SMN

2.7% positivity in myositis, Ag contributes to snRNPs assembly

Arthritis Rheum.2011 Jul;63（7）:1972-8.

cN1A

50% positivity for inclusion body myositisFront Immunol.2019 Apr 9;10:745.

10% positivity for MCTD, SSc or SLE overlap syndrome,
good steroid response, interstitial pneumonia is less common

Clinical Rheumatology and Related Research,
25：149〜158,2013

U2-RNP

Polymyositis-scleroderma overlap syndrome with less than
5% positivity, muscle weakness, and mild scleroderma

Clinical Rheumatology and Related Research,
25：149〜158,2013

Ku

Polymyositis-scleroderma overlap syndrome with 2−30%
positivity, muscle weakness, minimal skin sclerosis,
and good vital prognosis

Clinical Rheumatology and Related Research,
25：149〜158,2013

8−10% positivity in polymyositis-scleroderma overlap
syndrome, mild

Clinical Rheumatology and Related Research,
25：149〜158,2013

A positive rate of 1−2% in scleroderma,
Overlap of myositis and diﬀuse skin thickening occur frequently,
higher incidence in male and elderly patients

Arthritis Care Res（Hoboken）.
2014 Apr;66（4）:575-84.

10% positivity rate in overlap syndromes according
to each collagen disease.

Laboratory Medicine Encyclopedia & Dictionary

10−20% positivity for myositis

Laboratory Medicine Encyclopedia & Dictionary

Positive rate of 5% or less in myositis

Laboratory Medicine Encyclopedia & Dictionary

MJ（NXP-2）

U1-RNP̲70
U1-RNP̲A
U1-RNP̲C

PM-Scl１００
PM-Scl７５
RuvBL1/2
Ki
SS-A/Ro52
SS-A/Ro60
SS-B

6●Inquiries

Clinical Laboratory Center
307 Minatomachi, Marugame City, Kagawa 763-0042, Japan
TEL.0877-22-6276 FAX.0877-22-6280
E-mail th@fushimi.co.jp
HP address http://www.fushimi.co.jp/