人間 COPS5 ELISA Kit (HUFI03389)- 高感度

商品名:	Human COPS5 (COP9 signalosome complex subunit 5 ) ELISA Kit
製品コード:	HUFI03389
サイズ:	96T
エイリアス:	COP9 signalosome complex subunit 5 ELISA Kit, SGN5 ELISA Kit, Signalosome subunit 5 ELISA Kit, Jun activation domain-binding protein 1 ELISA Kit, CSN5 ELISA Kit, JAB1 ELISA Kit, COPS5 ELISA Kit
検出方法:	Sandwich ELISA, Double Antibody
申し込み:	This immunoassay kit allows for the in vitro quantitative determination of Human COPS5 (COP9 signalosome complex subunit 5 ) concentrations in serum plasma and other biological fluids.
感度:	< 18.75pg/ml
範囲:	31.25-2000pg/ml
保管所:	4°C for 6 months
ノート:	For Research Use Only

回復:

Matrices listed below were spiked with certain level of Human COPS5 (COP9 signalosome complex subunit 5 ) and the recovery rates were calculated by comparing the measured value to the expected amount of Human COPS5 (COP9 signalosome complex subunit 5 ) in samples.

Matrix	Recovery range(%)	Average(%)
serum(n=5)	86-102	96
EDTA plasma(n=5)	86-98	93
heparin plasma(n=5)	87-99	90

直線性:

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human COPS5 (COP9 signalosome complex subunit 5 ) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample	1:2	1:4	1:8
serum(n=5)	86-102%	89-103%	86-103%
EDTA plasma(n=5)	83-101%	88-97%	90-100%
UFH plasma(n=5)	81-88%	81-97%	82-96%

CV(%):

Intra-Assay: CV<8%
Inter-Assay: CV<10%

成分	量	保管所
ELISA Microplate (Dismountable)	8×12 strips	4°C for 6 months
Lyophilized Standard	2	4°C/-20°C
Sample/Standard Dilution Buffer	20ml	4°C
Biotin-labeled Antibody(Concentrated)	120ul	4°C (Protect from light)
Antibody Dilution Buffer	10ml	4°C
HRP-Streptavidin Conjugate(SABC)	120ul	4°C (Protect from light)
SABC Dilution Buffer	10ml	4°C
TMB Substrate	10ml	4°C (Protect from light)
Stop Solution	10ml	4°C
Wash Buffer(25X)	30ml	4°C
Plate Sealer	5	-

必要なその他の材料と設備:

Microplate reader with 450 nm wavelength filter
Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
Incubator
Deionized or distilled water
Absorbent paper
Buffer resevoir

UniProt Protein Function:	COPS5: Probable protease subunit of the COP9 signalosome complex (CSN), a complex involved in various cellular and developmental processes. The CSN complex is an essential regulator of the ubiquitin (Ubl) conjugation pathway by mediating the deneddylation of the cullin subunits of the SCF-type E3 ligase complexes, leading to decrease the Ubl ligase activity of SCF-type complexes such as SCF, CSA or DDB2. The complex is also involved in phosphorylation of p53/TP53, c-jun/JUN, IkappaBalpha/NFKBIA, ITPK1 and IRF8, possibly via its association with CK2 and PKD kinases. CSN-dependent phosphorylation of TP53 and JUN promotes and protects degradation by the Ubl system, respectively. In the complex, it probably acts as the catalytic center that mediates the cleavage of Nedd8 from cullins. It however has no metalloprotease activity by itself and requires the other subunits of the CSN complex. Interacts directly with a large number of proteins that are regulated by the CSN complex, confirming a key role in the complex. Component of the CSN complex, composed of COPS1/GPS1, COPS2, COPS3, COPS4, COPS5, COP6, COPS7 (COPS7A or COPS7B) and COPS8. In the complex, it probably interacts directly with COPS1, COPS2, COPS4, COPS6 and COPS7 (COPS7A or COPS7B). The CSN complex interacts with the BRISC complex. Also exists as monomeric form. Interacts with TP53, MIF, JUN, UCHL1, NCOA1, HIF1A, p27Kip1, BCL3, GFER, PGR, LHCGR, SMAD4, SMAD7, ID1, ID3, ITGB2 and TOP2A. Part of a complex consisting of RANBP9, Ran, DYRK1B and COPS5. Belongs to the peptidase M67A family. CSN5 subfamily.
UniProt Protein Details:	Protein type:Adaptor/scaffold; Translation; Protease; Transcription, coactivator/corepressor; Nuclear receptor co-regulator; EC 3.4.-.- Chromosomal Location of Human Ortholog: 8q13.1 Cellular Component: cytoplasm; eukaryotic translation initiation factor 3 complex; nucleoplasm; nucleus; signalosome; synaptic vesicle Molecular Function:metallopeptidase activity; protein binding; transcription coactivator activity; translation initiation factor activity; ubiquitin-specific protease activity Biological Process: cullin deneddylation; nucleotide-excision repair, DNA damage recognition; positive regulation of transcription from RNA polymerase II promoter; protein deneddylation; protein deubiquitination; regulation of JNK cascade; transcription from RNA polymerase II promoter; transcription-coupled nucleotide-excision repair; translation
NCBI Summary:	The protein encoded by this gene is one of the eight subunits of COP9 signalosome, a highly conserved protein complex that functions as an important regulator in multiple signaling pathways. The structure and function of COP9 signalosome is similar to that of the 19S regulatory particle of 26S proteasome. COP9 signalosome has been shown to interact with SCF-type E3 ubiquitin ligases and act as a positive regulator of E3 ubiquitin ligases. This protein is reported to be involved in the degradation of cyclin-dependent kinase inhibitor CDKN1B/p27Kip1. It is also known to be an coactivator that increases the specificity of JUN/AP1 transcription factors. [provided by RefSeq, Jul 2008]
UniProt Code:	Q92905
NCBI GenInfo Identifier:	55976562
NCBI Gene ID:	10987
NCBI Accession:	Q92905.4
UniProt Secondary Accession:	Q92905,O15386, Q6AW95, Q86WQ4, Q9BQ17,
UniProt Related Accession:	Q92905
Molecular Weight:	37,579 Da
NCBI Full Name:	COP9 signalosome complex subunit 5
NCBI Synonym Full Names:	COP9 signalosome subunit 5
NCBI Official Symbol:	COPS5
NCBI Official Synonym Symbols:	CSN5; JAB1; SGN5; MOV-34
NCBI Protein Information:	COP9 signalosome complex subunit 5
UniProt Protein Name:	COP9 signalosome complex subunit 5
UniProt Synonym Protein Names:	Jun activation domain-binding protein 1
UniProt Gene Name:	COPS5
UniProt Entry Name:	CSN5_HUMAN

*ノート: プロトコルは、各バッチ/ロットに固有です。正しい手順については、キットに含まれているプロトコルに従ってください。

ウェルに加える前に、SABCワーキング溶液とTMB基質を37°Cで少なくとも30分間平衡化します。サンプルと試薬を希釈するときは、完全に均一に混合する必要があります。各テストの標準曲線をプロットすることをお勧めします。

ステップ	プロトコル
1.	Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.	Aliquot 0.1ml standard solutions into the standard wells.
3.	Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4.	Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.	Seal the plate with a cover and incubate at 37 °C for 90 min.
6.	Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7.	Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.	Seal the plate with a cover and incubate at 37 °C for 60 min.
9.	Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10.	Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37 °C for 30 min.
11.	Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12.	Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37 °C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13.	Add 50 µL of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14.	Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

ELISAアッセイを実施する場合、可能な限り最良の結果を達成するためにサンプルを準備することが重要です。以下に、さまざまなサンプルタイプのサンプルを準備するための手順のリストを示します。

サンプルタイプ	プロトコル
血清	If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.
プラズマ	Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.
尿および脳脊髄液	Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
細胞培養上清	Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
細胞溶解物	Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
組織ホモジネート	The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.
組織溶解物	Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
母乳	Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

Antibodies

Anti-COPS5 Antibody (CAB13401)

Anti-COPS5 Antibody (CAB1766)

Anti-JAB1 Antibody (CAB4087)

COPS5 Antibody (PACO06593)

COPS5 Antibody (PACO14614)