Laminin alpha 4 Antibody [Unconjugated]

Western blot shows lysates of human placenta tissue. PVDF membrane was probed with 2 µg/mL of Sheep Anti-Human Laminin a4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7340) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for Laminin a4 at approximately 200-220 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Laminin a4 was detected in immersion fixed T98G human glioblastoma cell line, induced (upper panel) or not (lower panel) with StemXVivo EMT Inducing Media Supplement (Catalog # CCM017) , using Sheep Anti-Human Laminin a4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7340) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm and cell surfaces. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Laminin alpha 4 was detected in immersion fixed paraffin-embedded sections of human placenta using Sheep Anti-Human Laminin alpha 4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7340) at 3 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Sheep IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC006). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to Golgi apparatus. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Simple Western lane view shows lysates of human placenta tissue, loaded at 0.2 mg/mL. A specific band was detected for Laminin alpha 4 at approximately 233 kDa (as indicated) using 20 µg/mL of Sheep Anti-Human Laminin alpha 4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7340) followed by 1:50 dilution of HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). This experiment was conducted under reducing conditions and using the 66-440 kDa separation system.

Recombinant Human Laminin alpha 4 (Catalog # 7340-A4) supports adhesion in the HT1080 human fibrosarcoma cell line in a dose-dependent manner (orange line), as measured by Calcein AM. Adhesion elicited by Recombinant Human Laminin alpha 4 (5 µg/mL) is neutralized (green line) by increasing concentrations of Sheep Anti-Human Laminin alpha 4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7340). The ND50 is typically1.5-7.5 µg/mL.

Flow induces remodeling of the endothelial extracellular matrix.A, Proteome and (B) cell surface proteome data of identified laminins. T or CT represents the enzyme used for protein digestion. The depicted numbers represent the biotin-modified lysine residue. C, Schematic representation of laminin 411 with the linker region between domains LG3 and LG4. The blue lysine residues were found to be decreased under flow conditions (as depicted in panel B). D, Immunoblot analysis of LAMA4 in ECs exposed to flow over time. alpha -tubulin was used as a loading control. E, Protein distribution of LAMA5 and LAMA4 after flow-exposure. Green represents LAMA4 or LAMA5, blue is HOECHST. The scale bar is 10 μm. Depicted are maximum intensity projections. F, Proteome and (G) cell surface proteome data of identified fibulins. H, Immunofluorescence analysis of EFEMP1 and FBLN2 abundance and distribution affected by flow. Red is VE-cadherin, blue is HOECHST. Depicted are maximum intensity projections. Micrographs are representative for 3 independent experiments. The scale bar is 10 μm. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/32332107), licensed under a CC-BY license. Not internally tested by R&D Systems.

Knockdown of LAMA4 suppresses the malignant behaviour of osteosarcoma and accelerates ER stress-induced apoptosis. A The expression levels of LAMA4 in osteosarcoma (n = 88) and normal tissues (n = 564) were analysed from the TNMplot database. B LAMA4 was knocked down in MNNG/HOS and 143B cells. The expression of LAMA4 was detected by Western blot. GAPDH was used as the loading control. C-D The cells (3000 cells/well) with or without LAMA4 depletion were tested for cell growth in the colony formation assay. Viable colonies after 1 week were counted and are shown (C). Data are depicted as bar graphs (D). E-F The migration of the indicated cells was detected by Transwell assays. Representative images of crystal violet-stained culture plates are shown (E). Data are depicted as bar graphs (F). G-H MNNG/HOS cells with or without LAMA4 knockdown were treated with 3 μM TM for 36 h. Cell apoptosis and cell viability were analysed by Western blot (G) and CCK8 assays (H). GAPDH was used as the loading control. I-J MNNG/HOS cells with or without LAMA4 knockdown were treated with 1 μM TG for 36 h. Cell apoptosis and cell viability were analysed by Western blot (I) and CCK8 assays (J). GAPDH was used as the loading control. K-M MNNG/HOS cells (106 cells per mouse) with or without LAMA4 knockdown were injected subcutaneously into nude mice (n = 4). Representative images of xenograft tumours (K). The weights of the tumours were calculated and analysed (L). The expression levels of PARP, cleaved PARP and LAMA4 in tumours were measured by Western blot (M). Data in D, F, H, J, L were analysed by Student’s t test, *p 1 and q value 1 and q value 1 and q value 1 and q value 1 and q value 1 and q value < 0.05). The green dot indicates the downregulated genes, and the red dot indicates the upregulated genes. D The mRNA levels of LAMA4 are shown from RNA sequencing data. E-F The protein and mRNA levels of LAMA4 were detected by Western blot (E) and qRT–PCR (F) in MNNG/HOS cells with or without LINC00629 or KLF4 knockdown. Numbers represent the relative intensities of western blot bands of LAMA4 to GAPDH. G MNNG/HOS cells with or without KLF4 or LINC00629 knockdown were treated with 3 μM TM for 36 h. The expression of LAMA4 was detected by Western blot. Numbers represent the relative intensities of western blot bands of LAMA4 to GAPDH. H-I LINC00629 was overexpressed in MNNG/HOS cells with or without KLF4 knockdown. The protein and mRNA levels of LAMA4 were detected by Western blotting (H) and qRT–PCR (I). Numbers represent the relative intensities of western blot bands of LAMA4 to GAPDH. Data in F and M were analysed by Student’s t test, *p < 0.05, **p < 0.01, ***p < 0.001 Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/36539799), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Hu
• Applications: WB, Simple Western, IHC, B/N, ICC/IF
• Clonality: Polyclonal
• Host: Sheep
• Conjugate: Unconjugated
• Concentration: LYOPH

Laminin alpha 4 Antibody [Unconjugated] Summary

• Immunogen: Chinese hamster ovary cell line CHO-derived recombinant human Laminin alpha 4
  Gln826-Ala1816, predicted
  Accession # EAW48267
• Specificity: Detects human Laminin alpha 4 in direct ELISAs and Western blots. In direct ELISAs, approximately 15% cross-reactivity with recombinant mouse Laminin alpha 4 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Sheep
• Gene: LAMA4
• Purity Statement: Antigen Affinity-purified
• Endotoxin Note: <0.10 EU per 1 μg of the antibody by the LAL method.

Applications/Dilutions

• Dilutions:
  • Immunocytochemistry 5-15 ug/mL
  • Immunohistochemistry 3-25 ug/mL
  • Neutralization 1.5-7.5 ug/mL
  • Simple Western 20 ug/mL
  • Western Blot 2 ug/mL

Packaging, Storage & Formulations

• 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.
• Buffer: 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.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Sterile PBS to a final concentration of 0.2 mg/mL.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Laminin alpha 4 Antibody [Unconjugated]

• DKFZp686D23145
• LAMA3LAMA4*-1
• LAMA4
• laminin alpha 4 chain
• Laminin alpha 4
• laminin subunit alpha-4
• laminin, alpha 4
• Laminin-14 subunit alpha
• Laminin-8 subunit alpha
• Laminin-9 subunit alpha

Background

Laminin alpha 4 (LAMA-4 [Laminin subunit alpha 4]; also Laminin-14/423, -8/411 and -9/421 subunit alpha) is a 180-230 kDa secreted member of the laminin family of molecules. It is found in the basement membranes of adipocytes, endothelial cells, cardiac and visceral smooth muscle cells, fibroblasts, keratinocytes, and pancreatic acinar cells. Laminin is a heterotrimeric glycoprotein that attaches cells to basement membranes. It is composed of covalently-linked alpha -, beta - and gamma -chains, and is known to attach cells to basement membranes. In the case of the alpha -subunit, this is mediated through binding to multiple integrins ( alpha 3 beta 1, alpha v beta 3 and alpha 6 beta 1), plus dystroglycan and the syndecans-2 and -4. Mature human Laminin alpha 4 is 1799 amino acids (aa) in length (aa 25-1823). It contains multiple domains, including four EGF‑like domains (aa 82-255) and five Laminin G-like domains (aa 833-1820). There is a chondroitin sulfate attachment at the N‑terminus that accounts for 20-30 kDa of MW. Laminin alpha 4 is cleaved between the Laminin G-like 3 and 4 domains, creating a soluble 44 kDa fragment that possesses antibacterial activity. There are two potential splice variants, one that shows a deletion of aa 266-272, and another that contains a 55 aa substitution for aa 66-1823. Over aa 826-1816, human Laminin alpha 4 shares 91% aa sequence identity with mouse Laminin alpha 4.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for Laminin alpha 4 Antibody (AF7340)(9)

Publications using AF7340

• Hsieh, LT;Hall, BS;Newcombe, J;Mendum, TA;Varela, SS;Umrania, Y;Deery, MJ;Shi, WQ;Diaz-Delgado, J;Salguero, FJ;Simmonds, RE; The Mycobacterium ulcerans toxin mycolactone causes destructive Sec61-dependent loss of the endothelial glycocalyx and vessel basement membrane to drive skin necrosis eLife 2025-02-06 [PMID: 39913180] (Immunocytochemistry, Human)
• Saito-Diaz, K;Saini, T;Patel, AJ;James, C;Thomas, KS;Knight, TN;Gogita, SB;Zeltner, N; Laminin ?4 is required for the development of human peripheral sensory neurons bioRxiv : the preprint server for biology 2024-11-24 [PMID: 39605577] (Immunocytochemistry, Human)
• LT Hsieh, BS Hall, J Newcombe, TA Mendum, Y Umrania, MJ Deery, WQ Shi, FJ Salguero, RE Simmonds Mycolactone causes catastrophic Sec61-dependent loss of the endothelial glycocalyx and basement membrane: a new indirect mechanism driving tissue necrosis in Mycobacterium ulcerans infection bioRxiv : the preprint server for biology, 2023-02-21;0(0):. 2/21/2023 [PMID: 36865118]
• Yuan Wang, Shuo Zheng, Jian Han, Na Li, Renchen Ji, Xiaodong Li et al. LINC00629 protects osteosarcoma cell from ER stress-induced apoptosis and facilitates tumour progression by elevating KLF4 stability Journal of Experimental & Clinical Cancer Research 12/20/2022 [PMID: 36539799]
• B Zheng, J Qu, K Ohuchida, H Feng, SJF Chong, Z Yan, Y Piao, P Liu, N Sheng, D Eguchi, T Ohtsuka, K Mizumoto, Z Liu, S Pervaiz, P Gong, M Nakamura LAMA4 upregulation is associated with high liver metastasis potential and poor survival outcome of Pancreatic Cancer Theranostics, 2020-08-13;10(22):10274-10289. 2020-08-13 [PMID: 32929348] (Neutralization, Human)
• Eelke P. Béguin, Esmée F. J. Janssen, Mark Hoogenboezem, Alexander B. Meijer, Arie J. Hoogendijk, Maartje van den Biggelaar Flow-induced Reorganization of Laminin-integrin Networks Within the Endothelial Basement Membrane Uncovered by Proteomics Molecular & Cellular Proteomics 7/1/2020 [PMID: 32332107]
• Eelke P. Béguin, Esmée F. J. Janssen, Mark Hoogenboezem, Alexander B. Meijer, Arie J. Hoogendijk, Maartje van den Biggelaar Flow-induced Reorganization of Laminin-integrin Networks Within the Endothelial Basement Membrane Uncovered by Proteomics Molecular & Cellular Proteomics 2020-07-01 [PMID: 32332107] (Western Blot, Immunocytochemistry, Human)
• Y Yan, H Yu, L Sun, H Liu, C Wang, X Wei, F Song, H Li, H Ge, H Qian, X Li, X Tang, P Liu Laminin ?4 overexpression in the anterior lens capsule may contribute to the senescence of human lens epithelial cells in age-related cataract Aging (Albany NY), 2019-05-10;11(9):2699-2723. 2019-05-10 [PMID: 31076560] (Bioassay, Human)
• Ishikawa T, Wondimu Z, Oikawa Y, Ingerpuu S, Virtanen I, Patarroyo M Monoclonal antibodies to human laminin alpha4 chain globular domain inhibit tumor cell adhesion and migration on laminins 411 and 421, and binding of alpha6beta1 integrin and MCAM to alpha4-laminins. Matrix Biol, 2014-03-27;36(0):5-14. 2014-03-27 [PMID: 24681327] (Neutralization, Human)

Bioinformatics

• Gene Symbol: LAMA4
• Uniprot:
  • EAW48267()