Revision 5
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IHC-P, IF-IC

REACTIVITY:

Vir

SENSITIVITY:

Endogenous

MW (kDa):

110, 220

Source/Isotype:

Rabbit IgG

UniProt ID:

#P0DTC2

Entrez-Gene Id:

43740568

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunohistochemistry (Paraffin) 1:200 - 1:800
Immunofluorescence (Immunocytochemistry) 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb recognizes endogenous levels of total SARS-CoV-2 spike protein. The antibody detects full-length SARS-CoV-2 spike protein and also detects the S1 domain generated by furin cleavage. The antibody has been confirmed to detect the following SARS-CoV-2 isolates and variants: USA-WA1/2020, Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2), and Omicron (B.1.1.529). The antibody does not cross-react with spike proteins from SARS or MERS coronaviruses. Non-specific nuclear foci staining was observed in some mouse tissues by immunohistochemistry.

Species Reactivity:

Virus

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein corresponding to the S1 domain of SARS-CoV-2 spike protein (USA-WA1/2020 isolate).

Background

The cause of the COVID-19 pandemic is a novel and highly pathogenic coronavirus, termed SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2). SARS-CoV-2 is a member of the Coronaviridae family of viruses (1). The genome of SARS-CoV-2 is similar to other coronaviruses, and is comprised of four key structural proteins: S, the spike protein, E, the envelope protein, M, the membrane protein, and N, the nucleocapsid protein (2). Coronavirus spike proteins are class I fusion proteins and harbor an ectodomain, a transmembrane domain, and an intracellular tail (3,4). The highly glycosylated ectodomain projects from the viral envelope surface and facilitates attachment and fusion with the host cell plasma membrane. The ectodomain can be further subdivided into host receptor-binding domain (RBD) (S1) and membrane-fusion (S2) subunits, which are produced upon proteolysis by host proteases at S1/S2 and S2’ sites. S1 and S2 subunits remain associated after cleavage and assemble into crown-like homotrimers (2,4). In humans, both SARS-CoV and SARS-CoV-2 spike proteins utilize the angiotensin-converting enzyme 2 (ACE2) protein as a receptor for cellular entry (5-7). Spike protein subunits represent a key antigenic feature of coronavirus virions, and therefore represent an important target of vaccines, novel therapeutic antibodies, and small-molecule inhibitors (8,9).

  1. Zhou, P. et al. (2020) Nature 579, 270-3.
  2. Tortorici, M.A. and Veesler, D. (2019) Adv Virus Res 105, 93-116.
  3. Li, F. et al. (2006) J Virol 80, 6794-800.
  4. Li, F. (2016) Annu Rev Virol 3, 237-61.
  5. Shang, J. et al. (2020) Nature 581, 221-4.
  6. Wrapp, D. et al. (2020) Science 367, 1260-3.
  7. Yan, R. et al. (2020) Science 367, 1444-8.
  8. Yuan, Y. et al. (2017) Nat Commun 8, 15092.
  9. Amanat, F. and Krammer, F. (2020) Immunity 52, 583-9.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SignalStain is a registered trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品, (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Revision 5
#99423

SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb

Western Blotting Image 1: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb(上图)、Myc-Tag (71D10) Rabbit mAb #2278(中图)和 GAPDH (D16H11) XP® Rabbit mAb #5174(下图)对转染空载(泳道 1)或瞬时转染带血浆编码的 Myc/DDK 标记的 SARS-CoV-2 刺突蛋白(泳道 2)的 293T 细胞提取物进行蛋白质印迹分析。该抗体可检测全长(未裂解)SARS-CoV-2 刺突蛋白,以及与内源蛋白酶裂解产生的 S1 域相对应的片段。
Western Blotting Image 2: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb(上图)和 His-Tag (D3I1O) XP® Rabbit mAb #12698(下图)对 SARS-CoV-2 Spike S1-NTD (16-316) Recombinant Protein (8xHis-Tag) #88587(泳道 1)、SARS-CoV-2 Spike (trimeric) (16-1208) Recombinant Protein (8xHis-Tag) #65444(泳道 2)、SARS-CoV-2 Spike RBD (318-541) Recombinant Protein (8xHis-Tag) #48801(泳道 3)或 SARS-CoV-2 Spike RBD (multimeric) (319-591) Recombinant Protein (8xHis-Tag) #17862(泳道 4)进行蛋白质印迹分析。由于表位的位置(在 SARS-CoV-2 刺突蛋白的 S1 域内),该抗体检测到与 SARS CoV-2 刺突蛋白的全长胞外域和 S1-NTD 对应的重组蛋白,但未检测到仅对应于受体结合域 (RBD) 的重组蛋白。
Western Blotting Image 3: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb(上图)和 GAPDH (D16H11) XP® Rabbit mAb #5174(下图)对模拟感染的 Vero-E6 细胞(泳道 1)、SARS-CoV-2 感染的 Vero-E6 细胞(泳道 2)、分离的 SARS-CoV-2 病毒体(泳道 3)的提取物进行蛋白质印迹分析。该抗体可检测全长(未裂解)SARS-CoV-2 刺突蛋白,以及与内源蛋白酶裂解产生的 S1 域相对应的片段。SARS-CoV-2 病毒体和感染的 Vero-E6 细胞由波士顿大学国家新兴传染病实验室 Mohsan Saeed 博士提供。
Immunohistochemistry Image 1: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb 对石蜡包埋的 SARS-CoV-2 阳性人胎盘(左图,阳性)和正常人胎盘(右图,阴性)进行免疫组织化学分析。
Immunohistochemistry Image 2: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
以浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(右图)为对照,使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb(左图)对石蜡包埋的 SARS-CoV-2 感染的 hACE2 K18 小鼠肺细胞进行免疫组织化学分析。人ACE2 转基因小鼠肺组织由波士顿大学国家新兴传染病实验室 Nicholas Crossland 博士和 Florian Douam 博士友情提供。
Immunohistochemistry Image 3: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb 对石蜡包埋且转染带 SARS-CoV-2 刺突蛋白(左上图)的 293T 细胞沉淀物和以下各种石蜡包埋的人体组织进行免疫组织化学分析:结肠腺癌组织(右上图)、前列腺癌组织(右下图)和唾液腺小细胞癌组织(左下图)。注意组织中没有染色,所有染色均在 2019 之前完成,因此可作为可靠的阴性对照。
Immunohistochemistry Image 4: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb 对未转染(左图)或经过 SARS-CoV-2 刺突蛋白(右图)转染的石蜡包埋的 293T 细胞沉淀物进行免疫组织化学分析。
Immunohistochemistry Image 5: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
使用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb 对石蜡包埋的 A549-hACE2-hTMPRSS2 细胞沉淀物、模拟感染或感染的 SARS-CoV-2 2019-nCoV/USA-WA1/2020 变体、SARS-CoV-2 Alpha (B.1.1.7) 变体、SARS-CoV-2 Beta (B.1.351) 变体(如图所示,从左上图到右图)、SARS-CoV-2 Gamma (P.1) 变体、SARS-CoV-2 Delta (B.1.617.2) 变体或 SARS-CoV-2 Omicron (B.1.1.529) 变体(如图所示,从左下图到右图)进行免疫组织化学分析。A549-hACE2-hTMPRSS2 细胞沉淀物由波士顿大学国家新兴传染病实验室的 Nicholas Crossland 博士和 Florian Douam 博士慷慨提供。
Immunofluorescence Image 1: SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb Expand Image
对用 SARS-CoV-2 刺突蛋白瞬时转染、用 SARS-CoV-2 Spike Protein (S1) (E5S3V) Rabbit mAb(左图,绿色)标记以及用 DyLight 650 Phalloidin #12956(右图,红色)和 DAPI #4083(右图,蓝色)共同标记的 HCT 116 细胞进行共聚焦免疫荧光分析。