Mouse Clusterin DuoSet ELISA

R&D Systems | Catalog # DY2747

R&D Systems
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Key Product Details

Assay Type

Solid Phase Sandwich ELISA

Assay Range

0.469-30 ng/mL

Sample Type

Cell culture supernates, serum, and plasma
Note: Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet

Reactivity

Mouse

Mouse Clusterin DuoSet ELISA Features

  • Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
  • Development protocols are provided to guide further assay optimization
  • Assay can be customized to your specific needs
  • Economical alternative to complete kits
View all Clusterin ELISA Kits »

Product Summary for Mouse Clusterin DuoSet ELISA

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant mouse Clusterin. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

 

Product Specifications

Assay Format

96-well strip plate (sold separately)

Sample Volume Required

100 µL

Detection Method

Colorimetric ELISA - 450nm (TMB)

Conjugate

Biotin

Specificity

Please see the product datasheet

Label

HRP

Scientific Data Images for Mouse Clusterin DuoSet ELISA

Mouse Clusterin ELISA Standard Curve

Mouse Clusterin ELISA Standard Curve

Kit Contents for Mouse Clusterin DuoSet ELISA

  • Capture Antibody
  • Detection Antibody
  • Recombinant Standard
  • Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)

Other Reagents Required

DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.

The components listed above may be purchased separately:

PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or 0.05% Tween® 20 in PBS, pH 7.2-7.4

Reagent Diluent: (Catalog # DY995), or 1% BSA in PBS, pH 7.2-7.4, 0.2 µm filtered

Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)

Stop Solution: 2 N H2SO4 (Catalog # DY994)

Microplates: R&D Systems (Catalog # DY990)

Plate Sealers: ELISA Plate Sealers (Catalog # DY992)

Preparation and Storage

Shipping

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: Clusterin

Clusterin, also known as Apolipoprotein J, Sulfated Glycoprotein 2 (SGP-2), TRPM-2, and SP-40, is a secreted multifunctional 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 expressed in adult testis, ovary, adrenal gland, liver, heart, brain, and in many epithelial tissues during embryonic development (3). Mouse Clusterin is synthesized as a precursor that contains two coiled coil domains, two 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 approximately 80 kDa N-glycosylated and disulfide-linked heterodimer (7-9). Mature mouse Clusterin shares 77% and 93% amino acid sequence identity with human and rat Clusterin, respectively. 
An alternately spliced 50 kDa isoform of mouse Clusterin (nCLU) remains intracellular and is neither glycosylated nor cleaved into alpha and beta chains (4). Cellular exposure to ionizing radiation promotes the translocation of nCLU to the nucleus where it interacts with Ku70 and promotes apoptosis (4, 10). This function contrasts with the cytoprotective effect of secreted Clusterin (11). 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 (12, 13). The ability of Clusterin to bind and neutralize non-oxidatively modified LDL reduces cytotoxicity in atherosclerotic plaques (14). The chaperone function of Clusterin also helps to reduce the accumulation of beta -amyloid fibrils and damage due to amyloid plaques in Alzheimer's disease (15). Clusterin levels are elevated in the cerebrospinal fluid of patients with Alzheimer's disease, Parkinson's disease, and multiple sclerosis (9, 16, 17) and in the urine of patients with kidney injury or bladder cancer (18-20). Clusterin is released by activated platelets at sites of vascular injury and is found in the synovial fluid of rheumatoid arthritis and osteoarthritis patients (21, 22). During human tumor progression, nCLU is downregulated while the secreted form is upregulated and may be aberrantly glycosylated (10, 23-25). Increased circulating levels of Clusterin enhance tumor aggressiveness by inhibiting apoptosis and by promoting epithelial to mesenchymal transition (26-28).

Alternate Names

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

Entrez Gene IDs

1191 (Human); 12759 (Mouse)

Gene Symbol

CLU

Additional Clusterin Products

Product Documents for Mouse Clusterin DuoSet ELISA

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Mouse Clusterin DuoSet ELISA

For research use only

Citations for Mouse Clusterin DuoSet ELISA

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Protocols

View specific protocols for Mouse Clusterin DuoSet ELISA (DY2747):

GENERAL ELISA PROTOCOL

Plate Preparation

  1. Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
  2. Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
  3. Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
  4. Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.

Assay Procedure

  1. Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
  2. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  3. Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
  4. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  5. Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  6. Repeat the aspiration/wash as in step 2.
  7. Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  8. Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  9. Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.

 

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