Human HepaCAM Antibody

Catalog # Availability Size / Price Qty
MAB4108
MAB4108-SP

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Detection of Human HepaCAM by Western Blot.
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Product Details
Citations (9)
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Human HepaCAM Antibody Summary

Species Reactivity
Human
Specificity
Detects human HepaCAM in direct ELISAs and Western blots.
Source
Monoclonal Mouse IgG1 Clone # 419305
Purification
Protein A or G purified from hybridoma culture supernatant
Immunogen
Mouse myeloma cell line NS0-derived recombinant human HepaCAM
Val34-Tyr242
Accession # Q14CZ8
Formulation
Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
Label
Unconjugated

Applications

Recommended Concentration
Sample
Western Blot
0.25 µg/mL
See below

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Western Blot Detection of Human HepaCAM antibody by Western Blot. View Larger

Detection of Human HepaCAM by Western Blot. Western blot shows lysates of human brain (motor cortex) tissue and human brain (hippocampus) tissue. PVDF membrane was probed with 0.25 µg/mL of Mouse Anti-Human HepaCAM Monoclonal Antibody (Catalog # MAB4108) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for HepaCAM at approximately 45-70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Western Blot Detection of Human HepaCAM by Western Blot View Larger

Detection of Human HepaCAM by Western Blot HepaCAM associates with connexin 43.(A) U373 MG cells were stably transfected with pcDNA3.1 vector, wild-type hepaCAM, hepaCAM-R92Q and hepaCAM-R92W. Immunofluorescent staining was performed with antibodies against the hepaCAM cytoplasmic domain (green) and connexin 43 (red). Co-localization of hepaCAM and connexin 43 is indicated by yellow fluorescence. Nuclei were stained with DAPI (blue). Insets show a higher magnification of sites of cell-cell contacts. Cells were visualized by confocal microscopy under a 60× objective. Scale bar: 10 μm. (B) Co-immunoprecipitatation of connexin 43 and hepaCAM. Cell lysates were prepared from U373 MG cells stably transfected with pcDNA3.1 vector and wild-type hepaCAM, and immunoprecipitated with antibody against the hepaCAM extracellular domain (IP hepaCAM). Immunoprecipitation with mouse IgG1 (IP IgG) was included as a negative control. Western blot analysis was performed on the immunoprecipitates and input (3%) using connexin 43 antibody. The efficiency of hepaCAM immunoprecipitation was evaluated with an HRP-conjugated FLAG antibody. The IgG heavy chain detected with an HRP-conjugated anti-mouse antibody is shown as a loading control. (C) Co-immunoprecipitation of wild-type and mutant hepaCAM with connexin 43. Cell lysates were immunoprecipitated with antibody against the hepaCAM extracellular domain (IP hepaCAM). Immunoprecipitation with mouse IgG1 (IP IgG) was included as a negative control. Western blot analysis was performed on the immunoprecipitates and input (2%) using connexin 43 antibody. (D) Expression of wild-type hepaCAM increases connexin 43 protein levels in U373 MG cells. 20 μg of cell lysates were subjected to Western blot analysis. GAPDH was used as a loading control. The result presented is a representative experiment of four independent experiments with similar results. The full view blots are shown in Supplementary Figure 1. (E) Quantification of connexin 43 protein levels in D and in three additional independent Western blot analyses. Using ImageJ the densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands for each sample, and the mean relative density over the four experiments was calculated. The data presented are the means ± SE (n = 4), **p < 0.01 as assessed by one-way ANOVA with Tukey’s multiple comparison test. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep36218), licensed under a CC-BY license. Not internally tested by R&D Systems.

Western Blot Detection of Human HepaCAM by Western Blot View Larger

Detection of Human HepaCAM by Western Blot HepaCAM regulates connexin 43 stability.(A) Evaluation of connexin 43 mRNA expression. Total RNA was analyzed by RT-PCR. GAPDH and no template controls (NTC) were included as housekeeping gene and negative controls, respectively. (B) Evaluation of connexin 43 protein stability by a cycloheximide (CHX) chase assay. Cells treated with CHX (50 μg/ml) for the times indicated were lysed and 30 μg of cell lysates were subjected to Western blot analysis. The result presented is a representative experiment of three independent experiments with similar results. (C) Quantification of all three CHX chase experiments using ImageJ. The densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands at each time-point. The level of connexin 43 remaining at each time-point was calculated as a percentage of the initial connexin 43 level (time 0 of CHX treatment). The data presented are the means ± SE (n = 3). (D) Expression of hepaCAM in HEK293T cells increases connexin 43 protein levels. HEK293T cells were transiently transfected with pcDNA3.1 vector or wild-type hepaCAM. Two days after transfection, cells were lysed and 60 μg of cell lysates were subjected to Western blot analysis using antibodies against connexin 43 and the hepaCAM extracellular domain. The result presented is a representative experiment of three independent experiments with similar results. (E) Quantification of all three experiments using ImageJ. The densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands for each sample, and the mean relative density over the three experiments was calculated. The data presented are the means ± SE (n = 3), ***p < 0.0001 as assessed by t-test. (F) HepaCAM slows down connexin 43 turnover by the lysosomal pathway. U373 MG cells stably transfected with pcDNA3.1 vector or wild-type hepaCAM were treated with chloroquine (50 μM) and 30 μg of cell lysates were subjected to Western blot analysis for connexin 43. The result presented is representative of two independent experiments with similar results. The full view blots for (B,D,F) are shown in Supplementary Figures 3,4 and 5, respectively. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep36218), licensed under a CC-BY license. Not internally tested by R&D Systems.

Western Blot Detection of Human HepaCAM by Western Blot View Larger

Detection of Human HepaCAM by Western Blot HepaCAM regulates connexin 43 stability.(A) Evaluation of connexin 43 mRNA expression. Total RNA was analyzed by RT-PCR. GAPDH and no template controls (NTC) were included as housekeeping gene and negative controls, respectively. (B) Evaluation of connexin 43 protein stability by a cycloheximide (CHX) chase assay. Cells treated with CHX (50 μg/ml) for the times indicated were lysed and 30 μg of cell lysates were subjected to Western blot analysis. The result presented is a representative experiment of three independent experiments with similar results. (C) Quantification of all three CHX chase experiments using ImageJ. The densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands at each time-point. The level of connexin 43 remaining at each time-point was calculated as a percentage of the initial connexin 43 level (time 0 of CHX treatment). The data presented are the means ± SE (n = 3). (D) Expression of hepaCAM in HEK293T cells increases connexin 43 protein levels. HEK293T cells were transiently transfected with pcDNA3.1 vector or wild-type hepaCAM. Two days after transfection, cells were lysed and 60 μg of cell lysates were subjected to Western blot analysis using antibodies against connexin 43 and the hepaCAM extracellular domain. The result presented is a representative experiment of three independent experiments with similar results. (E) Quantification of all three experiments using ImageJ. The densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands for each sample, and the mean relative density over the three experiments was calculated. The data presented are the means ± SE (n = 3), ***p < 0.0001 as assessed by t-test. (F) HepaCAM slows down connexin 43 turnover by the lysosomal pathway. U373 MG cells stably transfected with pcDNA3.1 vector or wild-type hepaCAM were treated with chloroquine (50 μM) and 30 μg of cell lysates were subjected to Western blot analysis for connexin 43. The result presented is representative of two independent experiments with similar results. The full view blots for (B,D,F) are shown in Supplementary Figures 3,4 and 5, respectively. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep36218), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunocytochemistry/ Immunofluorescence Detection of Human HepaCAM by Immunocytochemistry/Immunofluorescence View Larger

Detection of Human HepaCAM by Immunocytochemistry/Immunofluorescence HepaCAM enhances connexin 43 expression.(A) Treatment of hepaCAM-expressing U373 MG cells with antibodies against the hepaCAM extracellular domain prevents the association of hepaCAM with connexin 43 at cell-cell contacts. Wild-type hepaCAM-expressing U373 MG cells were treated overnight with antibody against the hepaCAM extracellular domain in soluble form (10 μg/ml). Cells were also treated with the isotype mouse IgG1 as a control. The next day, cells were fixed and immunofluorescent staining was performed with antibodies against the hepaCAM extracellular domain (green) and connexin 43 (red). Co-localization of hepaCAM and connexin 43 is indicated by yellow fluorescence. Nuclei were stained with DAPI (blue). Cells were visualized by confocal microscopy under a 60× objective. Scale bar: 10 μm. (B) Treatment of hepaCAM-expressing U373 MG cells with antibodies against the hepaCAM extracellular domain causes a downregulation of connexin 43 expression. Wild-type hepaCAM-expressing U373 MG cells were treated overnight with antibody against the hepaCAM extracellular domain in soluble form (10 μg/ml). Cells were also treated with the isotype mouse IgG1 as a control. The next day, cells were lysed and 20 μg of cell lysates were subjected to Western blot analysis using connexin 43 antibody. GAPDH was used as a loading control. The full view blots are shown in Supplementary Figure 2. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep36218), licensed under a CC-BY license. Not internally tested by R&D Systems.

Immunocytochemistry/ Immunofluorescence Detection of Human HepaCAM by Immunocytochemistry/Immunofluorescence View Larger

Detection of Human HepaCAM by Immunocytochemistry/Immunofluorescence HepaCAM associates with connexin 43.(A) U373 MG cells were stably transfected with pcDNA3.1 vector, wild-type hepaCAM, hepaCAM-R92Q and hepaCAM-R92W. Immunofluorescent staining was performed with antibodies against the hepaCAM cytoplasmic domain (green) and connexin 43 (red). Co-localization of hepaCAM and connexin 43 is indicated by yellow fluorescence. Nuclei were stained with DAPI (blue). Insets show a higher magnification of sites of cell-cell contacts. Cells were visualized by confocal microscopy under a 60× objective. Scale bar: 10 μm. (B) Co-immunoprecipitatation of connexin 43 and hepaCAM. Cell lysates were prepared from U373 MG cells stably transfected with pcDNA3.1 vector and wild-type hepaCAM, and immunoprecipitated with antibody against the hepaCAM extracellular domain (IP hepaCAM). Immunoprecipitation with mouse IgG1 (IP IgG) was included as a negative control. Western blot analysis was performed on the immunoprecipitates and input (3%) using connexin 43 antibody. The efficiency of hepaCAM immunoprecipitation was evaluated with an HRP-conjugated FLAG antibody. The IgG heavy chain detected with an HRP-conjugated anti-mouse antibody is shown as a loading control. (C) Co-immunoprecipitation of wild-type and mutant hepaCAM with connexin 43. Cell lysates were immunoprecipitated with antibody against the hepaCAM extracellular domain (IP hepaCAM). Immunoprecipitation with mouse IgG1 (IP IgG) was included as a negative control. Western blot analysis was performed on the immunoprecipitates and input (2%) using connexin 43 antibody. (D) Expression of wild-type hepaCAM increases connexin 43 protein levels in U373 MG cells. 20 μg of cell lysates were subjected to Western blot analysis. GAPDH was used as a loading control. The result presented is a representative experiment of four independent experiments with similar results. The full view blots are shown in Supplementary Figure 1. (E) Quantification of connexin 43 protein levels in D and in three additional independent Western blot analyses. Using ImageJ the densities of the connexin 43 bands were normalized to the densities of the respective GAPDH bands for each sample, and the mean relative density over the four experiments was calculated. The data presented are the means ± SE (n = 4), **p < 0.01 as assessed by one-way ANOVA with Tukey’s multiple comparison test. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep36218), licensed under a CC-BY license. Not internally tested by R&D Systems.

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Preparation and Storage

Reconstitution
Reconstitute at 0.5 mg/mL in sterile PBS.
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Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: HepaCAM

HepaCAM (hepatocyte cell adhesion molecule) is a 50-75 kDa, type I transmembrane glycoprotein that belongs to the Ig-superfamily. It forms homodimers on the cell surface and promotes cell spreading and motility. Human HepaCAM is 382 aa in length. It contains a 206 aa extracellular domain (ECD) (aa 35‑240) and a 152 aa cytoplasmic region. The ECD has two C2 Ig-like domains. There is one potential truncated isoform that shows an alternate start site at Met200. Over aa 34‑242, human HepaCAM shares 99% and 98% aa sequence identity with mouse and dog HepaCAM, respectively.

Long Name
Hepatocyte Cell Adhesion Molecule
Entrez Gene IDs
220296 (Human)
Alternate Names
cancer susceptibility; FLJ25530; glial cell adhesion molecule; GlialCAM; HepaCAM; hepatic and glial cell adhesion molecule; hepatocyte cell adhesion molecule; Protein hepaCAM

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Citations for Human HepaCAM Antibody

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

9 Citations: Showing 1 - 9
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  1. Astroglial exosome HepaCAM signaling and ApoE antagonization coordinates early postnatal cortical pyramidal neuronal axon growth and dendritic spine formation
    Authors: Jin, S;Chen, X;Tian, Y;Jarvis, R;Promes, V;Yang, Y;
    Nature communications
    Species: Transgenic Mouse
    Sample Types: Whole Tissue
    Applications: IHC
  2. Identification of ligand-receptor pairs that drive human astrocyte development
    Authors: Voss, AJ;Lanjewar, SN;Sampson, MM;King, A;Hill, EJ;Sing, A;Sojka, C;Bhatia, TN;Spangle, JM;Sloan, SA;
    Nature neuroscience
    Species: Human
    Sample Types: Organoid
    Applications: Immunopanning
  3. Lymphocyte deficiency alters the transcriptomes of oligodendrocytes, but not astrocytes or microglia
    Authors: MC Krawczyk, L Pan, AJ Zhang, Y Zhang
    PLoS ONE, 2023-02-24;18(2):e0279736.
    Species: Mouse
    Sample Types: Whole Cells
    Applications: Immunoprecipitation
  4. Mutations in the transcriptional regulator MeCP2 severely impact key cellular and molecular signatures of human astrocytes during maturation
    Authors: J Sun, S Osenberg, A Irwin, LH Ma, N Lee, Y Xiang, F Li, YW Wan, IH Park, M Maletic-Sa, N Ballas
    Cell Reports, 2023-01-05;42(1):111942.
    Species: Human
    Sample Types: Organoid
    Applications: IHC
  5. Enrichment of Glial Cells From Human Post-mortem Tissue for Transcriptome and Proteome Analysis Using Immunopanning
    Authors: A Nolle, I van Dijken, CM Waelti, D Calini, J Bryois, E Lezan, S Golling, A Augustin, L Foo, JJM Hoozemans
    Frontiers in Cellular Neuroscience, 2021-12-13;15(0):772011.
    Species: Human
    Sample Types: Whole Cells
    Applications: Immunopanning
  6. Neurotoxic reactive astrocytes induce cell death via saturated lipids
    Authors: KA Guttenplan, MK Weigel, P Prakash, PR Wijewardha, P Hasel, U Rufen-Blan, AE Münch, JA Blum, J Fine, MC Neal, KD Bruce, AD Gitler, G Chopra, SA Liddelow, BA Barres
    Nature, 2021-10-06;0(0):.
    Species: Mouse
    Sample Types: Whole Cells
    Applications: Bioassay
  7. Knockout of reactive astrocyte activating factors slows disease progression in an ALS mouse model
    Authors: KA Guttenplan, MK Weigel, DI Adler, J Couthouis, SA Liddelow, AD Gitler, BA Barres
    Nat Commun, 2020-07-27;11(1):3753.
    Species: Mouse
    Sample Types: Whole Cells
    Applications: Cell Culture
  8. Tumor-associated reactive astrocytes aid the evolution of immunosuppressive environment in glioblastoma
    Authors: D Henrik Hei, VM Ravi, SP Behringer, JH Frenking, J Wurm, K Joseph, NWC Garrelfs, J Strähle, S Heynckes, J Grauvogel, P Franco, I Mader, M Schneider, AL Potthoff, D Delev, UG Hofmann, C Fung, J Beck, R Sankowski, M Prinz, O Schnell
    Nat Commun, 2019-06-11;10(1):2541.
    Species: Human
    Sample Types: Whole Cells, Whole Tissue
    Applications: Flow Cytometry, IHC, Immunopanning
  9. HepaCAM associates with connexin 43 and enhances its localization in cellular junctions
    Sci Rep, 2016-11-07;6(0):36218.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot

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