CXCL10 ELISA Kit

Catalog No. ABIN625033

Product Details CXCL10 ELISA Kit

Target: CXCL10 (Chemokine (C-X-C Motif) Ligand 10 (CXCL10))

Reactivity: Human

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 8-6000 pg/mL

Minimum Detection Limit: 8 pg/mL

Application: ELISA

Purpose: Human IP-10 (CXCL10) ELISA Kit for cell culture supernatants, plasma, and serum samples.

Sample Type: Plasma, Cell Culture Supernatant, Serum

Analytical Method: Quantitative

Specificity: This ELISA kit shows no cross-reactivity with any of the cytokines tested: Human BDNF, BLC, ENA-78, FGF-4, IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-4, IL-5, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12 p70, IL-12 p40, IL-13, IL-15, I-309, G-CSF, GM-CSF, IFN-gamma, Leptin (OB), MCP-1, MCP- 2, MCP-3, MDC, MIP-1 alpha, MIP-1 beta, MIP-1 delta, PARC, PDGF, RANTES, SCF, TARC, TGF-beta, TIMP-1, TIMP-2, TNF-alpha, TNF-beta, TPO, VEGF.

Sensitivity: < 8 pg/mL

Characteristics:

• Strip plates and additional reagents allow for use in multiple experiments
• Quantitative protein detection
• Establishes normal range
• The best products for confirmation of antibody array data

Components:

• Pre-Coated 96-well Strip Microplate
• Wash Buffer
• Stop Solution
• Assay Diluent(s)
• Lyophilized Standard
• Biotinylated Detection Antibody
• Streptavidin-Conjugated HRP
• TMB One-Step Substrate

Material not included:

• Distilled or deionized water
• Precision pipettes to deliver 2 μL to 1 μL volumes
• Adjustable 1-25 μL pipettes for reagent preparation
• 100 μL and 1 liter graduated cylinders
• Tubes to prepare standard and sample dilutions
• Absorbent paper
• Microplate reader capable of measuring absorbance at 450nm
• Log-log graph paper or computer and software for ELISA data analysis

Application Details

Application Notes: Recommended Dilution for serum and plasma samples2 fold

Sample Volume: 100 μL

Plate: Pre-coated

Protocol:

1. Prepare all reagents, samples and standards as instructed in the manual.
2. Add 100 μL of standard or sample to each well.
3. Incubate 2.5 h at RT or O/N at 4 °C.
4. Add 100 μL of prepared biotin antibody to each well.
5. Incubate 1 h at RT.
6. Add 100 μL of prepared Streptavidin solution to each well.
7. Incubate 45 min at RT.
8. Add 100 μL of TMB One-Step Substrate Reagent to each well.
9. Incubate 30 min at RT.
10. Add 50 μL of Stop Solution to each well.
11. Read at 450 nm immediately.

Reagent Preparation:

1. Bring all reagents and samples to room temperature (18-25 °C) before use.
   2. Sample dilution: If your samples need to be diluted, Assay Diluent A (Item D) should be used for dilution of serum/plasma samples. 1x Assay Diluent B (Item E) should be used for dilution of culture supernatants and urine. Suggested dilution for normal serum/plasma: 2 fold. Please note that levels of the target protein may vary between different specimens. Optimal dilution factors for each sample must be determined by the investigator.
   3. Assay Diluent B should be diluted 5-fold with deionized or distilled water.
   4. Preparation of standard: Briefly spin the vial of Item C. Add 400 µL Assay Diluent A (for serum/plasma samples) or 1x Assay Diluent B (for cell culture medium and urine) into Item C vial to prepare a 50 ng/mL standard. Dissolve the powder thoroughly by a gentle mix. Add 80 µL IP-10 standard from the vial of Item C, into a tube with 586.7 µL Assay Diluent A or 1x Assay Diluent B to prepare a 6000 pg/mL stock standard solution. Pipette 400 µL Assay Diluent A or 1x Assay Diluent B into each tube. Use the stock standard solution to produce a dilution series . Mix each tube thoroughly before the next transfer. Assay Diluent A or 1x Assay Diluent B serves as the zero standard (0 pg/mL). 200 µL 80 µL standard +586.7 µL 200myl 200 µL 200 µL 200 µL 200 µL 6000 2000 666.7 222.2 74.07 24.69 8.23 0 pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL
   5. If the Wash Concentrate (20x) (Item B) contains visible crystals, warm to room temperature and mix gently until dissolved. Dilute 20 ml of Wash Buffer Concentrate into deionized or distilled water to yield 400 ml of 1x Wash Buffer.
   6. Briefly spin the Detection Antibody vial (Item F) before use. Add 100 µL of 1x Assay Diluent B into the vial to prepare a detection antibody concentrate. Pipette up and down to mix gently (the concentrate can be stored at 4 °C for 5 days). The detection antibody concentrate should be diluted 100-fold with 1x Assay Diluent B and used in step 4 of Part VI Assay Procedure.
   7. Briefly spin the HRP-Streptavidin concentrate vial (Item G) and pipette up and down to mix gently before use. HRP-Streptavidin concentrate should be diluted 500-fold with 1x Assay Diluent B. For example: Briefly spin the vial (Item G) and pipette up and down to mix gently . Add 30 µL of HRP-Streptavidin concentrate into a tube with 15 ml 1x Assay Diluent B to prepare a final 500 fold diluted HRP-Streptavidin solution (don't store the diluted solution for next day use). Mix well.

Assay Procedure:

1. Bring all reagents and samples to room temperature (18 - 25 °C) before use. It is recommended that all standards and samples be run at least in duplicate.
   2. Add 100 µL of each standard (see Reagent Preparation step 2) and sample into appropriate wells. Cover well and incubate for 2.5 hours at room temperature or over night at 4 °C with gentle shaking.
   3. Discard the solution and wash 4 times with 1x Wash Solution. Wash by filling each well with Wash Buffer (300 myl) using a multi-channel Pipette or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
   4. Add 100 µL of 1x prepared biotinylated antibody (Reagent Preparation step 6) to each well. Incubate for 1 hour at room temperature with gentle shaking.
   5. Discard the solution. Repeat the wash as in step
   6. Add 100 µL of prepared Streptavidin solution (see Reagent Preparation step 7) to each well. Incubate for 45 minutes at room temperature with gentle shaking.
   7. Discard the solution. Repeat the wash as in step
   8. Add 100 µL of TMB One-Step Substrate Reagent (Item H) to each well. Incubate for 30 minutes at room temperature in the dark with gentle shaking.
   9. Add 50 µL of Stop Solution (Item I) to each well. Read at 450 nm immediately.

Calculation of Results:

Calculate the mean absorbance for each set of duplicate standards, controls and samples, and subtract the average zero standard optical density. Plot the standard curve on log-log graph paper or using Sigma plot software, with standard concentration on the x-axis and absorbance on the y-axis. Draw the best-fit straight line through the standard points.
Typical Data: These standard curves are for demonstration only. A standard curve must be run with each assay. Assay Diluent A IP-10 concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10 Assay Diluent B IP-10 concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10
Sensitivity: The minimum detectable dose of IP-10 is typically less than 8 pg/mL.
Recovery: Recovery was determined by spiking various levels of human IP-10 into human serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 94.49 82-104 Plasma 96.13 83-103 Cell culture media 101.34 84-105
Linearity: Sample Type Serum Plasma Cell culture media 1:2 Average % of Expected 92 94 98 Range ( %) 82-103 84-104 86-105 1:4 Average % of Expected 97 96 95 Range ( %) 83-105 84-104 82-103
Reproducibility: Intra-Assay: CV<10 % Inter-Assay: CV<12 %

Assay Precision: Intra-Assay: CV< 10 % Inter-Assay: CV< 12 %

Restrictions: For Research Use only

Handling

Handling Advice: Avoid repeated freeze-thaw cycles.

Storage: -20 °C

Storage Comment: The entire kit may be stored at -20°C for up to 1 year from the date of shipment. Avoid repeated freeze-thaw cycles. The kit may be stored at 4°C for up to 6 months. For extended storage, it is recommended to store at -80°C.

Expiry Date: 6 months

References for CXCL10 ELISA Kit (ABIN625033)

Persidsky, Hill, Zhang, Dykstra, Winfield, Reichenbach, Potula, Mukherjee, Ramirez, Rom: "Dysfunction of brain pericytes in chronic neuroinflammation." in: Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism, Vol. 36, Issue 4, pp. 794-807, (2016) (PubMed).

Spaks, Jaunalksne, Spaka, Chudasama, Pirtnieks, Krievins: "Diagnostic Value of Circulating CXC Chemokines in Non-small Cell Lung Cancer." in: Anticancer research, Vol. 35, Issue 12, pp. 6979-83, (2016) (PubMed).

Gao, Camous, Lu, Lim, Larbi, Ng: "Novel inflammatory markers associated with cognitive performance: Singapore Longitudinal Ageing Studies." in: Neurobiology of aging, Vol. 39, pp. 140-6, (2016) (PubMed).

Marozin, Altomonte, Muñoz-Álvarez, Rizzani, De Toni, Thasler, Schmid, Ebert: "STAT3 inhibition reduces toxicity of oncolytic VSV and provides a potentially synergistic combination therapy for hepatocellular carcinoma." in: Cancer gene therapy, Vol. 22, Issue 6, pp. 317-25, (2016) (PubMed).

Ko, Kuo, Chang, Chen, Liu, Chen, Tsai, Lee, Chen, Wu, Chen: "CXCL10/IP-10 is a biomarker and mediator for Kawasaki disease." in: Circulation research, Vol. 116, Issue 5, pp. 876-83, (2015) (PubMed).

Gunluoglu, Seyhan, Kazancioglu, Gunluoglu, Veske, Yazar, Altin: "Diagnosing latent tuberculosis in immunocompromised patients measuring blood IP-10 production capacity: an analysis of chronic renal failure patients." in: Internal medicine (Tokyo, Japan), Vol. 54, Issue 5, pp. 465-72, (2015) (PubMed).

Edvardsen, Bjånesøy, Hellesen, Breivik, Bakke, Husebye, Bratland: "Peripheral Blood Cells from Patients with Autoimmune Addison's Disease Poorly Respond to Interferons In Vitro, Despite Elevated Serum Levels of Interferon-Inducible Chemokines." in: Journal of interferon & cytokine research : the official journal of the International Society for Interferon and Cytokine Research, Vol. 35, Issue 10, pp. 759-70, (2015) (PubMed).

Wu, Lu, Wang, Jin, Cheng, Fang, Wang, Ma, Zhang, Cheng: "Clinical symptoms, immune factors, and molecular characteristics of an adult male in Shenzhen, China infected with influenza virus H5N1." in: Journal of medical virology, Vol. 85, Issue 5, pp. 760-8, (2013) (PubMed).

Hardy, Sieg, Rodriguez, Anthony, Asaad, Jiang, Mudd, Schacker, Funderburg, Pilch-Cooper, Debernardo, Rabin, Lederman, Harding: "Interferon-? is the primary plasma type-I IFN in HIV-1 infection and correlates with immune activation and disease markers." in: PLoS ONE, Vol. 8, Issue 2, pp. e56527, (2013) (PubMed).

Bratland, Hellesen, Husebye: "Induction of CXCL10 chemokine in adrenocortical cells by stimulation through toll-like receptor 3." in: Molecular and cellular endocrinology, Vol. 365, Issue 1, pp. 75-83, (2013) (PubMed).

Fujiwara, Nagai, Oniki, Yoshimoto, Nishigori: "Interleukin (IL)-17 versus IL-27: opposite effects on tumor necrosis factor-?-mediated chemokine production in human keratinocytes." in: Experimental dermatology, Vol. 21, Issue 1, pp. 70-2, (2011) (PubMed).

Cakir, Levendoglu, Kiyici, Coskun: "Serum CXCL10 levels and neuromuscular manifestations in patients with autoimmune thyroid diseases." in: Autoimmunity, Vol. 44, Issue 6, pp. 496-503, (2011) (PubMed).

Vargas-Inchaustegui, Hogg, Tulliano, Llanos-Cuentas, Arevalo, Endsley, Soong: "CXCL10 production by human monocytes in response to Leishmania braziliensis infection." in: Infection and immunity, Vol. 78, Issue 1, pp. 301-8, (2010) (PubMed).

Angiolillo, Sgadari, Taub, Liao, Farber, Maheshwari, Kleinman, Reaman, Tosato: "Human interferon-inducible protein 10 is a potent inhibitor of angiogenesis in vivo." in: The Journal of experimental medicine, Vol. 182, Issue 1, pp. 155-62, (1995) (PubMed).

Luster, Leder: "IP-10, a -C-X-C- chemokine, elicits a potent thymus-dependent antitumor response in vivo." in: The Journal of experimental medicine, Vol. 178, Issue 3, pp. 1057-65, (1993) (PubMed).

Gottlieb, Luster, Posnett, Carter: "Detection of a gamma interferon-induced protein IP-10 in psoriatic plaques." in: The Journal of experimental medicine, Vol. 168, Issue 3, pp. 941-8, (1988) (PubMed).

Daniel, Smith, Barnum: "The relationship of California mastitis test (C.M.T.) scores with leucocyte counts on bucket milk samples." in: The Canadian veterinary journal. La revue vétérinaire canadienne, Vol. 7, Issue 4, pp. 80-3 (PubMed).

Target Details for CXCL10

Target: CXCL10 (Chemokine (C-X-C Motif) Ligand 10 (CXCL10))

Alternative Name: IP-10 (CXCL10 Products)

Synonyms: CXCL10 ELISA Kit, C7 ELISA Kit, IFI10 ELISA Kit, INP10 ELISA Kit, IP-10 ELISA Kit, SCYB10 ELISA Kit, crg-2 ELISA Kit, gIP-10 ELISA Kit, mob-1 ELISA Kit, CRG-2 ELISA Kit, IP10 ELISA Kit, Ifi10 ELISA Kit, Scyb10 ELISA Kit, C-X-C motif chemokine ligand 10 ELISA Kit, C-X-C motif chemokine 10 ELISA Kit, chemokine (C-X-C motif) ligand 10 ELISA Kit, CXCL10 ELISA Kit, cxl10 ELISA Kit, Cxcl10 ELISA Kit

Background: IP-10 (Interferon-inducible protein-10) is also called gamma-IP-10 or INP-10. It has a length of 98 amino acids and belongs to the family of chemotactic cytokines. IP-10 has been detected in keratinocytes, lymphocytes, monocytes, and endothelial cells in immunologically mediated processes. It has been suggested that IP-10 may play an important role in hypersensitivity reactions of the delayed type. Increased levels of IP-10 are found in psoriatic plaques characterized by the infiltration of neutrophils. The Human IP-10 ELISA (Enzyme-Linked Immunosorbent Assay) kit is an in vitro enzyme-linked immunosorbent assay for the quantitative measurement of human IP-10 in serum, plasma, cell culture supernatants and urine. This assay employs an antibody specific for human IP-10 coated on a 96-well plate. Standards and samples are pipetted into the wells and IP-10 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-human IP-10 antibody is added. After washing away unbound biotinylated antibody, HRP-conjugated streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of IP-10 bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm. Reproducibility: Intra-Assay: CV<10% Inter-Assay: CV<12%.

Gene ID: 3627

UniProt: P02778