Human SCD (SCD1) knockout HeLa cell line (ab265220)
製品の概要
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製品名
Human SCD (SCD1) knockout HeLa cell line -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, 4 bp deletion in exon 3 and Insertion of the selection cassette in exon 3 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
アプリケーション
適用あり: WBmore details -
Biosafety level
2 -
特記事項
Recommended control: Human wild-type HeLa cell line (ab255928). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: DMEM (High Glucose) + 10% FBS
Initial handling guidelines: Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method. Based on cell count, seed cells in an appropriate cell culture flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules.
4. Incubate the culture at 37°C incubator with 5% CO2. Cultures should be monitored daily.Subculture guidelines:
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
We will provide viable cells that proliferate on revival.
製品の特性
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Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Cervix -
Cell type
epithelial -
Disease
Adenocarcinoma -
Gender
Female -
STR Analysis
Amelogenin X D5S818: 11, 12 D13S317: 12, 13.3 D7S820: 8, 12 D16S539: 9, 10 vWA: 16, 18 TH01: 7 TPOX: 8,12 CSF1PO: 9, 10 -
Mycoplasma free
Yes -
保存方法
Shipped on Dry Ice. Store in liquid nitrogen. -
バッファー
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
研究分野
ターゲット情報
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機能
Terminal component of the liver microsomal stearyl-CoA desaturase system, that utilizes O(2) and electrons from reduced cytochrome b5 to catalyze the insertion of a double bond into a spectrum of fatty acyl-CoA substrates including palmitoyl-CoA and stearoyl-CoA. -
配列類似性
Belongs to the fatty acid desaturase family. -
ドメイン
The histidine box domains may contain the active site and/or be involved in metal ion binding. -
細胞内局在
Endoplasmic reticulum membrane. - Information by UniProt
関連製品
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KO cell lysates
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Related Products
アプリケーション
The Abpromise guarantee
Abpromise保証は、 次のテスト済みアプリケーションにおけるab265220の使用に適用されます
アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション | Abreviews | 特記事項 |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 42 kDa.
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特記事項 |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 42 kDa. |
画像
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All lanes : Anti-SCD1 antibody [CD.E10] (ab19862) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : SCD knockout HeLa cell lysate
Lane 3 : HepG2 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 42 kDa
Observed band size: 36 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab19862 observed at 36 kDa. Red - loading control ab2866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55kDa.
ab19862 was shown to react with SCD1 in HeLa wild-type cells in western blot with loss of signal observed in SCD knockout cell line ab265220 (SCD knockout cell lysate ab257658). HeLa wild-type and SCD knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab19862 and ab2866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4°C at a 1 in 1000 Dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-SCD1 antibody [EPR21963] (ab236868) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : SCD knockout HeLa cell lysate
Lane 3 : HepG2 cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution
Predicted band size: 42 kDaanes 1-3: Merged signal (red and green). Green - ab236868. Red - loading control ab8245 observed at 50 kDa.
ab236868 Anti-SCD1 antibody [EPR21963] was shown to specifically react with SCD1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265220 (knockout cell lysate ab257658) was used. Wild-type and SCD1 knockout samples were subjected to SDS-PAGE. ab236868 and Anti-tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.
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Allele-1: 4 bp deletion in exon 3.
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Allele-2: 4 bp deletion in exon 3.
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Allele-3: 4 bp deletion in exon 3.
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Allele-5: Insertion of the selection cassette in exon 3.
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Allele-4: Insertion of the selection cassette in exon 3.
プロトコール
データシートおよび資料
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SDS download
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Datasheet download
参考文献 (1)
ab265220 は 1 報の論文で使用されています。
- Yin H et al. HIF-1a downregulation of miR-433-3p in adipocyte-derived exosomes contributes to NPC progression via targeting SCD1. Cancer Sci 112:1457-1470 (2021). PubMed: 33511729