Hamster CXCL10/IP-10(Interferon Gamma Induced Protein 10kDa) ELISA Kit - Information
|Hamster CXCL10/IP-10(Interferon Gamma Induced Protein 10kDa) ELISA Kit (HMFI0014-C)||Tissue, Cell Culture Supernatant, Cell Lysate||Competitive|
|Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit (HMFI0014-S)||Serum/Plasma||Sandwich|
The Assay Genie Hamster CXCL10/IP-10(Interferon Gamma Induced Protein 10kDa) ELISA Kit can assay for CXCL10 in the following samples: tissue, cell culture supernatant, cell Lysate.
How our Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit works?
The Assay Genie ELISA (enzyme-linked immunosorbent assays) assay kits are designed for the quantitative measurement of analytes in a wide variety of samples. As today's scientists demand high quality consistent data for high impact journals, Assay Genie have developed a range of sensitive, fast and reliable ELISA kit assays to meet and exceed those demands. Our assay kits use a quantitative sandwich ELISA technique and each kit comes with highly specific antibodies pre-coated onto a 96-well microtiter plate.
At Assay Genie we understand the need for speed! Therefore, we have developed an ultra-fast protocol meaning you achieve your results rapidly. So, once you have prepared and plated your samples, blanks and standards, you simply incubate with a highly specific biotin-conjugated primary antibody and Avidin conjugated to Horseradish Peroxidase (HRP) and incubate for the appropriate length of time. After washing the plate according to the protocol and addition of the TMB (3,3',5,5'-Tetramethylbenzidine) solution, the appearance of a blue colour should be detected due to an enzymatic reaction catalysed by HRP. Next step is the addition of the Stop Solution which terminates the HRP reaction and the blue colour turns yellow with the signal intensity measured on a plate reader at 450nm. The amount of bound Pentraxin 3 / TSG-14 is proportional to the signal generated by the reaction meaning the kit assay gives you a quantitative measurement of the analyte in your samples.
Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit - Data
|Product Sku|| |
C-X-C motif chemokine ELISA Kit, IP10 ELISA Kit, Cxcl10 ELISA Kit, Interferon Gamma Induced Protein 10kDa ELISA Kit, IP-10 ELISA Kit
|Detection method|| |
2-8'C for 6 months
Matrices listed below were spiked with certain level of CXCL10 and the recovery rates were calculated by comparing the measured value to the expected amount of CXCL10 in samples.
The linearity of the kit was assayed by testing samples spiked with appropriate concentration of CXCL10 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
For Research Use Only
Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit Protocol
The below protocol is a sample protocol for Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit using a biotinylated detection antibody and streptavidin-HRP. Sandwich ELISAs allow for the detection and quantification of an analyte in a sample by using known analyte concentrations as standards and plotting absorbance of known concentrations vs known standard concentrations. This allows the researcher to calculate the amount of CXCL10 present in their sample.
When diluting samples and reagents, they must be mixed completely and evenly. Before adding TMB into wells, equilibrate TMB Substrate for 30 minutes at 37°C. It is recommended to plot a standard curve for each test.
|1.||Set standard, test samples (diluted at least 1/2 with Sample Dilution Buffer), control (blank) wells on the pre-coated plate respectively, and then, records their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (blank) wells!|
|2.||Prepare Standards: Aliquot 100ul of zero tube, 1st tube, 2nd tube, 3rd tube, 4th tube, 5th tube, 6th tube and Sample Dilution Buffer (blank) into the standard wells|
|3.||Add Samples: Add 100ul of properly diluted sample into test sample wells.|
|4.||Incubate: Seal the plate with a cover and incubate at 37°C for 90 minutes.|
|5.||Wash: Remove the cover and discard the plate content, and wash plate 2 times with Wash Buffer. Do NOT let the wells dry completely at any time.|
|6.||Biotin-labeled Antibody: Add 100ul Biotin-labeled antibody working solution into above wells (standard, test sample and blank wells). Add the solution at the bottom of each well without touching the sidewall, cover the plate and incubate at 37°C for 60 minutes.|
|7.||Wash: Remove the cover, and wash plate 3 times with Wash Buffer, and let the Wash Buffer stay in the wells for 1-2 minutes each time|
|8.||HRP-Streptavidin Conjugate (SABC): Add 100ul of SABC Working Solution into each well, cover the plate and incubate at 37°C for 30 minutes|
|9.||Wash: Remove the cover and wash plate 5 times with Wash Buffer, and let the wash buffer stay in the wells for 1-2 minutes each time.|
|10.||TMB Substrate: Add 90ul TMB Substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 minutes. (Note: The reaction time can be shortened or extended according to the actual color change, but not more than 30 minutes. You can terminate the reaction when apparent gradient appeared in standard wells.)|
|11.||Stop: Add 50ul Stop Solution into each well. The color will turn yellow immediately. The adding order of Stop Solution should be as the same as the TMB Substrate Solution.|
|12.||OD Measurement: Read the O.D. absorbance at 450nm in Microplate Reader immediately after adding the stop solution.|
Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit components
|ELISA Microplate(Dismountable)||8×12 strips||-20°C|
|Sample/Standard Dilution Buffer||20ml||2-8°C|
|Biotin-labeled Antibody(Concentrated)||120ul||2-8°C (Protect from light)|
|Antibody Dilution Buffer||10ml||2-8°C|
|HRP-Streptavidin Conjugate(SABC)||120ul||2-8°C (Protect from light)|
|SABC Dilution Buffer||10ml||2-8°C|
|TMB Substrate||10ml||2-8°C (Protect from light)|
Other materials and equipment required:
The Assay Genie Hamster CXCL10/IP-10 (Interferon Gamma Induced Protein 10kDa) ELISA Kit will require other equipment and materials to carry out the assay. Please see list below for further details.
- Microplate reader with 450 nm wavelength filter
- Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
- Deionized or distilled water
- Absorbent paper
- Buffer resevoir
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
Place whole blood sample at room temperature for 2 hours or put it at 2-8°C overnight and centrifugation for 20 minutes at approximately 1000×g, Collect the supernatant and carry out the assay immediately. Blood collection tubes should be disposable, non-pyrogenic, and non-endotoxin.
Collect plasma using EDTA-Na2 or heparin as an anticoagulant. Centrifuge samples for 15 minutes at 1000×g at 2-8°C within 30 minutes of collection. Collect the supernatant and carry out the assay immediately. Avoid hemolysis, high cholesterol samples.
As hemolysis blood has relation to assay result, it is necessary to remove residual blood by washing tissue with pre-cooling PBS buffer (0.01M, pH=7.4). Mince tissue after weighing it and get it homogenized in PBS (the volume depends on the weight of the tissue. Normal, 9mL PBS would be appropriate to 1 gram tissue pieces. Some protease inhibitors are recommended to add into the PBS) with a glass homogenizer on ice. To further break the cells, you can sonicate the suspension with an ultrasonic cell disrupter or subject it to freeze-thaw cycles. The homogenates are then centrifuged for 5 minutes at 5000×g to get the supernatant. The total protein concentration was determined by BCA kit and the total protein concentration of each pore sample should not exceed 0.3mg.
Cell culture supernatant
Centrifuge supernatant for 20 minutes at 1000×g at 2-8°C to remove insoluble impurity and cell debris. Collect the clear supernatant and carry out the assay immediately.
Cell Culture Lysate
Commercial RIPA kits are recommended to follow the instructions provided. Generally, 0.5ml RIPA lysis buffer would be appropriate to 2x106 cells, DNA must to be removed. The total protein concentration was determined by BCA kit and the total protein concentration of each pore sample should not exceed 0.3mg.
Other Biological Fluids
Centrifuge samples for 20 minutes at 1000×g at 2-8°C. Collect supernatant and carry out the assay immediately.