Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human

Cited:

Human

Applications

Validated:

Immunohistochemistry, Western Blot, Immunoprecipitation

Cited:

Western Blot, Immunocytochemistry, Immunoprecipitation

Label

Unconjugated

Antibody Source

Polyclonal Goat IgG
View all Clusterin Primary Antibodies »

Product Specifications

Immunogen

Mouse myeloma cell line NS0-derived recombinant human Clusterin isoform 1
Asp75-Glu501
Accession # NP_001822

Specificity

Detects human Clusterin in direct ELISAs and Western blots. In direct ELISAs and Western blots, approximately 40% cross-reactivity with recombinant mouse Clusterin is observed and less than 5% cross-reactivity with recombinant rat Clusterin is observed.

Clonality

Polyclonal

Host

Goat

Isotype

IgG

Scientific Data Images for Human Clusterin Isoform 1 Antibody

Detection of Human Clusterin antibody by Western Blot.

Detection of Human Clusterin by Western Blot.

Western blot shows lysates of human liver tissue and human serum. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human Clusterin Isoform 1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2937) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). Specific bands were detected for Clusterin Precusor at approximately 60-65 kDa and Clusterin a and beta chains at approximately 36 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Clusterin antibody in Human Alzheimer's Disease Brain by Immunohistochemistry (IHC-P).

Clusterin in Human Alzheimer's Disease Brain.

Clusterin was detected in immersion fixed paraffin-embedded of human Alzheimer's disease brain using Human Clusterin Isoform 1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2937) at 5 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.
Detection of Human Clusterin/APOJ by Western Blot

Detection of Human Clusterin/APOJ by Western Blot

Extracellular clusterin interacts with alpha ‐synuclein preformed fibrils (pffs). (a) Medium from primary astrocytes treated with alpha ‐synuclein pffs for 16 hr, or with monomeric alpha ‐synuclein (m), were subjected to immunoblotting using clusterin antibody. Ponceau‐S staining was used as a loading control. (b) Quantification of clusterin is normalized to GAPDH protein of cell lysates. Data are from eight independent experiments and are expressed as the mean ± SD. Data were analyzed using unpaired t test; **p =.0028. (c) alpha ‐Synuclein pffs isolated by centrifugation from total medium of primary astrocytes treated with alpha ‐synuclein pffs, or monomeric alpha ‐synuclein (M), for 16 hr were subjected to immunoblotting using clusterin and alpha ‐synuclein antibodies. Asterisk in the alpha ‐synuclein immunoblot indicates clusterin bands still present after Clusterin antibody stripping. (d) Clusterin immunoprecipitated from the medium of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr was subjected to immunoblotting using an alpha ‐synuclein antibody. (e) Quantification of clusterin— alpha ‐synuclein pffs interaction has been obtained by normalization of alpha ‐synuclein pffs bound to clusterin for alpha ‐synuclein pffs bound to protein‐G beads. Data are representative of three independent experiments and are expressed as the mean ± SD. Data were analyzed using one‐sample t test; *p = .0493. (f) Z‐stack frames of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (green), cadherin (red), and nuclei with DAPI (blue). Scale bar 10 μm. (g) Maximum intensity Z‐projection confocal images of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (red), Early Endosome Antigen 1 (EEA1) (green) and nuclei with DAPI (blue). Scale bar 10 μm. Individual points of the graphs represent each single experiment [Color figure can be viewed at wileyonlinelibrary.com] Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/33045109), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Human Clusterin/APOJ by Western Blot

Detection of Human Clusterin/APOJ by Western Blot

Extracellular clusterin interacts with alpha ‐synuclein preformed fibrils (pffs). (a) Medium from primary astrocytes treated with alpha ‐synuclein pffs for 16 hr, or with monomeric alpha ‐synuclein (m), were subjected to immunoblotting using clusterin antibody. Ponceau‐S staining was used as a loading control. (b) Quantification of clusterin is normalized to GAPDH protein of cell lysates. Data are from eight independent experiments and are expressed as the mean ± SD. Data were analyzed using unpaired t test; **p =.0028. (c) alpha ‐Synuclein pffs isolated by centrifugation from total medium of primary astrocytes treated with alpha ‐synuclein pffs, or monomeric alpha ‐synuclein (M), for 16 hr were subjected to immunoblotting using clusterin and alpha ‐synuclein antibodies. Asterisk in the alpha ‐synuclein immunoblot indicates clusterin bands still present after Clusterin antibody stripping. (d) Clusterin immunoprecipitated from the medium of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr was subjected to immunoblotting using an alpha ‐synuclein antibody. (e) Quantification of clusterin— alpha ‐synuclein pffs interaction has been obtained by normalization of alpha ‐synuclein pffs bound to clusterin for alpha ‐synuclein pffs bound to protein‐G beads. Data are representative of three independent experiments and are expressed as the mean ± SD. Data were analyzed using one‐sample t test; *p = .0493. (f) Z‐stack frames of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (green), cadherin (red), and nuclei with DAPI (blue). Scale bar 10 μm. (g) Maximum intensity Z‐projection confocal images of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (red), Early Endosome Antigen 1 (EEA1) (green) and nuclei with DAPI (blue). Scale bar 10 μm. Individual points of the graphs represent each single experiment [Color figure can be viewed at wileyonlinelibrary.com] Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/33045109), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Human Clusterin/APOJ by Western Blot

Detection of Human Clusterin/APOJ by Western Blot

Extracellular clusterin interacts with alpha ‐synuclein preformed fibrils (pffs). (a) Medium from primary astrocytes treated with alpha ‐synuclein pffs for 16 hr, or with monomeric alpha ‐synuclein (m), were subjected to immunoblotting using clusterin antibody. Ponceau‐S staining was used as a loading control. (b) Quantification of clusterin is normalized to GAPDH protein of cell lysates. Data are from eight independent experiments and are expressed as the mean ± SD. Data were analyzed using unpaired t test; **p =.0028. (c) alpha ‐Synuclein pffs isolated by centrifugation from total medium of primary astrocytes treated with alpha ‐synuclein pffs, or monomeric alpha ‐synuclein (M), for 16 hr were subjected to immunoblotting using clusterin and alpha ‐synuclein antibodies. Asterisk in the alpha ‐synuclein immunoblot indicates clusterin bands still present after Clusterin antibody stripping. (d) Clusterin immunoprecipitated from the medium of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr was subjected to immunoblotting using an alpha ‐synuclein antibody. (e) Quantification of clusterin— alpha ‐synuclein pffs interaction has been obtained by normalization of alpha ‐synuclein pffs bound to clusterin for alpha ‐synuclein pffs bound to protein‐G beads. Data are representative of three independent experiments and are expressed as the mean ± SD. Data were analyzed using one‐sample t test; *p = .0493. (f) Z‐stack frames of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (green), cadherin (red), and nuclei with DAPI (blue). Scale bar 10 μm. (g) Maximum intensity Z‐projection confocal images of primary astrocytes treated with alpha ‐synuclein pffs for 16 hr and stained for alpha ‐synuclein (purple), clusterin (red), Early Endosome Antigen 1 (EEA1) (green) and nuclei with DAPI (blue). Scale bar 10 μm. Individual points of the graphs represent each single experiment [Color figure can be viewed at wileyonlinelibrary.com] Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/33045109), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Rat Clusterin/APOJ by Western Blot

Detection of Rat Clusterin/APOJ by Western Blot

Identification of a mitochondrial CLU protein isoform.(A) DIV 9 Mitotracker-stained (green) primary neurons were probed for CLU immunoreactivity using anti-CLU H-330 (red) and visualized using 40X confocal microscopy. (B) Pure cortical mitochondria were isolated as indicated. Equal concentrations of whole tissue lysate, crude mitochondria, and pure mitochondria were analyzed via SDS-PAGE and probed for CLU immunoreactivity using anti-CLU H-330 (left panel) and anti-CLU M-18 (right panel) (n = 3 independent isolations). Biochemical characterization of isolated fractions was performed using a panel of organelle-specific antibodies: voltage-dependent anion channels (VDAC) (mitochondria), calnexin and calreticulin (ER), and Gapdh (cytosol). (C) Crude and pure mitochondria were isolated and subjected to endoglycosidase treatment using PNGase F and Endo H. Deglycosylated mitochondrial lysates were then analyzed for CLU immunoreactivity using anti-CLU M-18. Red font: deglycosylated protein isoforms; blue font: isoforms that were unaffected by glycosidase treatment; green asterisk: excess Endo H enzyme.Positive controls for deglycosylation studies.RNase B, a high mannose glycoprotein, has a single N-linked glycosylation site and was used as a positive control for endoglycosidases that cleave N-linked carbohydrates. Fetuin, a glycoprotein containing sialylated N-linked and O-linked glycans, was used as a positive control for endoglycosidases that cleave both N-linked and O-linked carbohydrates. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/31738162), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Rat Clusterin/APOJ by Western Blot

Detection of Rat Clusterin/APOJ by Western Blot

Identification of a mitochondrial CLU protein isoform.(A) DIV 9 Mitotracker-stained (green) primary neurons were probed for CLU immunoreactivity using anti-CLU H-330 (red) and visualized using 40X confocal microscopy. (B) Pure cortical mitochondria were isolated as indicated. Equal concentrations of whole tissue lysate, crude mitochondria, and pure mitochondria were analyzed via SDS-PAGE and probed for CLU immunoreactivity using anti-CLU H-330 (left panel) and anti-CLU M-18 (right panel) (n = 3 independent isolations). Biochemical characterization of isolated fractions was performed using a panel of organelle-specific antibodies: voltage-dependent anion channels (VDAC) (mitochondria), calnexin and calreticulin (ER), and Gapdh (cytosol). (C) Crude and pure mitochondria were isolated and subjected to endoglycosidase treatment using PNGase F and Endo H. Deglycosylated mitochondrial lysates were then analyzed for CLU immunoreactivity using anti-CLU M-18. Red font: deglycosylated protein isoforms; blue font: isoforms that were unaffected by glycosidase treatment; green asterisk: excess Endo H enzyme.Positive controls for deglycosylation studies.RNase B, a high mannose glycoprotein, has a single N-linked glycosylation site and was used as a positive control for endoglycosidases that cleave N-linked carbohydrates. Fetuin, a glycoprotein containing sialylated N-linked and O-linked glycans, was used as a positive control for endoglycosidases that cleave both N-linked and O-linked carbohydrates. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/31738162), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for Human Clusterin Isoform 1 Antibody

Application
Recommended Usage

Immunohistochemistry

5-15 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human Alzheimer's disease brain

Immunoprecipitation

25 µg/mL
Sample: Conditioned cell culture medium spiked with Recombinant Human Clusterin (Catalog # 2937‑HS), see our available Western blot detection antibodies

Western Blot

1 µg/mL
Sample: Human liver tissue and human serum

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

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.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

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.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: Clusterin

Clusterin, also known as Apolipoprotein J, Sulfated Glycoprotein 2 (SGP-2), TRPM-2, and SP-40,40, is a secreted multi-functional protein that was named for its ability to induce cellular clustering. It binds a wide range of molecules and may function as a chaperone of misfolded extracellular proteins. It also participates in the control of cell proliferation, apoptosis, and carcinogenesis (1, 2). Clusterin is predominantly expressed in adult testis, ovary, adrenal gland, liver, heart, and brain and in many epithelial tissues during embryonic development (3). Human Clusterin is synthesized as a precursor that contains two coiled coil domains, three nuclear localization signals (NLS), and one heparin binding domain (4-6). Intracellular cleavages of the precursor remove the signal peptide and generate comparably sized alpha and beta chains which are secreted as an 80 kDa N-glycosylated disulfide-linked heterodimer (7, 8). Mature human Clusterin shares 77% amino acid sequence identity with mouse and rat Clusterin. High μg/mL concentrations of Clusterin circulate predominantly as a component of high density lipoprotein particles, and these are internalized and degraded through interactions with LRP-2/Megalin (9, 10). In human, an alternately spliced 50 kDa isoform of Clusterin (nCLU) lacks the signal peptide and remains intracellular (5, 11). This molecule is neither glycosylated nor cleaved into alpha and beta chains (11). In the cytoplasm, nCLU destabilizes the actin cytoskeleton and inhibits NF kappa B activation (12, 13). Cellular exposure to ionizing radiation promotes the translocation of nCLU to the nucleus where it interacts with Ku70 and promotes apoptosis (5, 11). This function contrasts with the cytoprotective effect of secreted Clusterin (14). During colon cancer tumor progression there is a downregulation of the intracellular form and an upregulation of the glycosylated secreted form (11).

References

  1. Carver, J.A. et al. (2003) IUBMB Life 55:661.
  2. Shannan, B. et al. (2006) Cell Death Differ. 13:12.
  3. French, L.E. et al. (1993) J. Cell Biol. 122:1119.
  4. Kirszbaum, L. et al. (1989) EMBO J. 8:711.
  5. Leskov, K.S. et al. (2003) J. Biol. Chem. 278:11590.
  6. Pankhurst, G.J. et al. (1998) Biochemistry 37:4823.
  7. Burkey, B.F. et al. (1991) J. Lipid. Res. 32:1039.
  8. de Silva, H.V. et al. (1990) J. Biol. Chem. 265:14292.
  9. Jenne, D.E. et al. (1991) J. Biol. Chem. 266:11030.
  10. Kounnas, M.Z. et al. (1995) J. Biol. Chem. 270:13070.
  11. Pucci, S. et al. (2004) Oncogene 23:2298.
  12. Moretti, R.M. et al. (2007) Cancer Res. 67:10325.
  13. Santilli, G. et al. (2003) J. Biol. Chem. 278:38214.
  14. Trougakos, I.P. et al. (2004) Cancer Res. 64:1834.

Alternate Names

APOJ, CLI, CLU, SGP-2, SP-40, TRPM-2

Entrez Gene IDs

1191 (Human); 12759 (Mouse)

Gene Symbol

CLU

UniProt

NP_001822

Additional Clusterin Products

Product Documents for Human Clusterin Isoform 1 Antibody

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Product Specific Notices for Human Clusterin Isoform 1 Antibody

For research use only

Citations for Human Clusterin Isoform 1 Antibody

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