Anti-CD74抗体(ab64772)
Key features and details
- Rabbit polyclonal to CD74
- Suitable for: WB, ICC/IF, IP, IHC-P
- Knockout validated
- Reacts with: Human
- Isotype: IgG
选择批间可重复性更高的重组抗体
- 研究可靠 —— 各批次间结果一致且可重复
- 长期批量供应 —— 采用重组技术,可实现快速生产
- 首次实验即可成功 —— 经过大量验证确认了特异性
- 符合伦理标准 —— 产品不含动物成分
概述
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产品名称
Anti-CD74抗体
参阅全部 CD74 一抗 -
描述
兔多克隆抗体to CD74 -
宿主
Rabbit -
经测试应用
适用于: WB, ICC/IF, IP, IHC-Pmore details -
种属反应性
与反应: Human
预测可用于: Non human primates -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- WB: Raji and Daudi cell lysates. IF/ICC: Raw246.7 cell line. IHC-P: Human tonsil and lymph node tissues. IP: Raji whole cell extract.
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常规说明
The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.
If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle. -
存储溶液
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help. -
Concentration information loading...
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纯度
Immunogen affinity purified -
克隆
多克隆 -
同种型
IgG -
研究领域
相关产品
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
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Recombinant Protein
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab64772于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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WB |
Use a concentration of 1 µg/ml. Detects a band of approximately 34 kDa (predicted molecular weight: 34 kDa).
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ICC/IF |
Use a concentration of 1 µg/ml.
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IP |
Use a concentration of 5 µg/ml.
IP image from Phase V. Lot JLD 04.06.2013 |
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IHC-P | (1) |
1/80 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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说明 |
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WB
Use a concentration of 1 µg/ml. Detects a band of approximately 34 kDa (predicted molecular weight: 34 kDa). |
ICC/IF
Use a concentration of 1 µg/ml. |
IP
Use a concentration of 5 µg/ml. IP image from Phase V. Lot JLD 04.06.2013 |
IHC-P
1/80 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. |
靶标
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功能
Plays a critical role in MHC class II antigen processing by stabilizing peptide-free class II alpha/beta heterodimers in a complex soon after their synthesis and directing transport of the complex from the endoplasmic reticulum to the endosomal/lysosomal system where the antigen processing and binding of antigenic peptides to MHC class II takes place. Serves as cell surface receptor for the cytokine MIF. -
序列相似性
Contains 1 thyroglobulin type-1 domain. -
细胞定位
Cell membrane. Endoplasmic reticulum membrane. Golgi apparatus > trans-Golgi network. Endosome. Lysosome. Transits through a number of intracellular compartments in the endocytic pathway. It can either undergo proteolysis or reach the cell membrane. - Information by UniProt
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数据库链接
- Entrez Gene: 972 Human
- Omim: 142790 Human
- SwissProt: P04233 Human
- Unigene: 436568 Human
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别名
- CD 74 antibody
- CD74 antibody
- CD74 antigen (invariant polypeptide of major histocompatibility complex, class II antigen-associated) antibody
see all
图片
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All lanes : Anti-CD74 antibody (ab64772) at 1 µg/ml
Lane 1 : Wild-type Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Lane 2 : CD74 knockout Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Lane 3 : Daudi (Human Burkitt's lymphoma cell line) whole cell lysate
Lane 4 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 34 kDa
Observed band size: 35 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab64772 observed at 35 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab64772 was shown to react with CD74 in wild-type Raji cells in Western blot with loss of signal observed in CD74 knockout cell line ab273876 (knockout cell lysate ab273830). Wild-type Raji and CD74 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab64772 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at 1 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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ab64772 (1:160) staining CD74 in paraffin-embedded human tonsil (left panel) using an automated system (Ventana Discovery). Right-hand panel shows negative control (no primary antibody).
Using this protocol there is strong membrane staining of activated B cells in the germinal centres and B cells of the mantle zone of the follicles plus scattered cells of the interfollicular areas (paracortical B cells).Sections were rehydrated and antigen retrieved in CC1 Cell Conditioning Buffer using Ventana Mild Retrieval programme. Slides were blocked in 3% H2O2 / 4 min / 37°C and incubated with ab64772 (1:160 dilution / 2 hours / 37°C). Sections then blocked (4mins / 37°C) and incubated with Dako swine anti-rabbit antibody (1:50, 28 min / 37°C). Staining was amplified and detected by incubation with Ventana Streptavidin ABC system (16 min / 37°C) and Ventana DAB map reagent (8 min / 37°C). Slides were counterstained with Haematox
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CD74 was immunoprecipitated using 0.5mg Raji whole cell extract, 5µg of Rabbit polyclonal to CD74 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, Raji whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab64772.
Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
Band: 34kDa; CD74
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All lanes : Anti-CD74 antibody (ab64772) at 1 µg/ml
Lane 1 : Wild-type Raji cell lysate
Lane 2 : CD74 CRISPR/Cas9 edited Raji cell lysate
Lane 3 : Jurkat cell lysate
Lane 4 : HepG2 cell lysate
Lysates/proteins at 30 µg per lane.
Performed under reducing conditions.
Predicted band size: 34 kDa
Observed band size: 35 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab64772 observed at 35 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab64772 was shown to react with CD74 in western blot. The band observed in CD74 CRISPR/Cas9 edited cell line ab273378 (CRISPR/Cas9 edited lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab64772 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at 1 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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Anti-CD74 antibody (ab64772) at 1 µg/ml + Raji (Human Burkitt's lymphoma cell line) Whole Cell Lysate at 10 µg
Secondary
Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Performed under reducing conditions.
Predicted band size: 34 kDa
Observed band size: 34 kDa
Exposure time: 2 minutes -
ab64772 (1:80) staining CD74 in paraffin-embedded human lymph node (left panel) using an automated system (Ventana Discovery). Right-hand panel shows negative control (no primary antibody).
Using this protocol there is strong membrane staining of activated B cells in the germinal centres and B cells of the mantle zone of the follicles plus scattered cells of the interfollicular areas (paracortical B cells).Sections were rehydrated and antigen retrieved in CC1 Cell Conditioning Buffer using Ventana Mild Retrieval programme. Slides were blocked in 3% H2O2 / 4 min / 37°C and incubated with ab64772 (1:80 dilution / 1 hour / 37°C). Sections then blocked (3mins / 37°C) and incubated with Dako swine anti-rabbit antibody (1:50, 28 min / 37°C). Staining was amplified and detected by incubation with Ventana Streptavidin ABC system (16 min / 37°C) and Ventana DAB map reagent (8 min / 37°C). Slides were counterstained with Haematoxyli
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ab64772 (1/250) staining CD74 in paraffin-embedded Human tonsil tissue. Tissue underwent fixation in formaldehyde, peroxidase blocking, protein blocking and heat mediated antigen retrieval. The secondary antibody was goat anti rabbit conjugated to HRP. For further experimental details please refer to abreview.
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ICC/IF image of ab64772 stained RAW246.7 cells. The cells were 4% paraformaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab64772, 1µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
实验方案
数据表及文件
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SDS download
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Datasheet download
文献 (4)
ab64772 被引用在 4 文献中.
- O'Connor MH et al. A follicular regulatory Innate Lymphoid Cell population impairs interactions between germinal center Tfh and B cells. Commun Biol 4:563 (2021). PubMed: 33980982
- Graham A & Nothnick WB Concurrent Immunohistochemical Localization and Western Blot Analysis of the MIF Receptor, CD74, in Formalin-Fixed, Paraffin-Embedded Tissue. Methods Mol Biol 2080:123-134 (2020). PubMed: 31745876
- Swanson MEV et al. Quantitative immunohistochemical analysis of myeloid cell marker expression in human cortex captures microglia heterogeneity with anatomical context. Sci Rep 10:11693 (2020). PubMed: 32678124
- Nothnick WB et al. Macrophage Migration Inhibitory Factor Receptor, CD74, is Overexpressed in Human and Baboon ( Papio Anubis) Endometriotic Lesions and Modulates Endometriotic Epithelial Cell Survival and Interleukin 8 Expression. Reprod Sci 25:1557-1566 (2018). PubMed: 29592775