SOCS1 ELISA Kit

Produktnummer ABIN578750

Produktdetails SOCS1 ELISA Kit

Target: SOCS1 (Suppressor of Cytokine Signaling 1 (SOCS1))

Reaktivität: Human

Nachweismethode: Colorimetric

Detektionsbereich: 0.156-10 ng/mL

Untere Nachweisgrenze: 0.156 ng/mL

Applikation: ELISA

Verwendungszweck: This immunoassay kit allows for the in vitro quantitative determination of human beta- glucuronidase,GUS concentrations in human tissue or cell culture supernates.

Proben: Cell Culture Supernatant, Cell Culture Supernatant

Analytische Methode: Quantitative

Spezifität: This assay recognizes recombinant and natural human GUS.

Kreuzreaktivität (Details): No significant cross-reactivity or interference was observed.

Sensitivität: The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest detectable concentration that could be differentiated from zero.

Produktmerkmale: Homo sapiens,Human,Suppressor of cytokine signaling 1,SOCS-1,JAK-binding protein,JAB,STAT-induced STAT inhibitor 1,SSI-1,Tec-interacting protein 3,TIP-3,SOCS1,SSI1,TIP3

Bestandteile: Reagent (Quantity ): Assay plate (1×20ml), Standard (2), Sample Diluent (1 × 20ml), Assay Diluent A (1x10ml), Assay Diluent B (1x10ml), Detection Reagent A (1 × 120μl), Detection Reagent B (1 × 120μl), Wash Buffer (25 x concentrate) (1 × 30ml), Substrate (1x10ml), Stop Solution (1x10ml), Plate sealer for 96 wells (5), Instruction (1)

Benötigtes Material: Luminometer. Pipettes and pipette tips. EP tube Deionized or distilled water.

Anwendungsinformationen

Probenmenge: 100 μL

Plattentyp: Pre-coated

Protokoll: beta -Glucuronidase is a glycosylhydrolase commonly found in animals, plants and microrganisms. The physiological importance of this enzyme in the metabolism of sulfated glycosaminoglycans is well known, and its genetic deficiency in humans causes a mucopolysaccharide storage disease known as mucopolysaccharidosis type VII. On the other hand, molecular characterization of beta -glucuronidase has unequivocally established the structure of the active site by X-ray crystallography and by site-directed mutagenesis.Based on the comparison of amino acid sequences in the active sites, most beta -glucuronidases, which have been identified in human, dog, cat, rat, mouse and E.coli, are classified as family 2 glycosylhydrolases with two conserved motifs (family 2 glycosylhydrolase signature and family 2 glycosylhydrolase acid-base catalyst).

Aufbereitung der Reagenzien:

Bring all reagents to room temperature before use. Wash Buffer - If crystals have formed in the concentrate, warm to room temperature and mix gently until the crystals have completely dissolved. Dilute 30 mL of Wash Buffer Concentrate into deionized or distilled water to prepare 750 mL of Wash Buffer. Standard - Reconstitute the Standard with 1.0 mL of Sample Diluent. This reconstitution 3 produces a stock solution of 400 ng/mL. Allow the standard to sit for a minimum of 15 minutes with gentle agitation prior to making serial dilutions (Making serial dilution in the wells directly is not permitted). Please firstly dilute the stock solution to 100 ng/mL and the diluted standard serves as the high standard (100 ng/mL). The Sample Diluent serves as the zero standard (0 ng/mL). ng/mL 400 100 50 25 12.5 6.25 3.12 1.56 0 Detection Reagent A and B - Dilute to the working concentration using Assay Diluent A and B (1:100), respectively.

Probennahme: 1. Grind fresh tissue to a powder with liquid nitrogen in a mortar and pestle. 2. Add 3 times volume of samples extraction buffer(10%TCA) at -20 overnight. C 3. Centrifuge at 8000rpm for 1h at 4 to collect precipitated protein,decant supernatant. C 4. Add the same volume of ice cold 100% acetone, centrifuge at 8000rpm for 15min at 4 ,then dry vacuum deposition in reserve. C 5. Add lysis buffer(2.7g urea,0.2g CHAPS add dH20 to 5ml) place at room temperature for 30 minutes,then centrifuge at 8000rpm for 15min at 4, store samples at 4 provisional. C C Note: Tissue homogenates and cell culture supernatant samples to be used within 7 days may be stored at 2-8 C, otherwise samples must stored at -20 C ( ≤ 1 months) or -80 C ( ≤ 2 months) to avoid loss of bioactivity and contamination. Avoid freeze-thaw cycles. When performing the assay slowly bring samples to room temperature.

Testdurchführung:

Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37 C directly.). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at 4 C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their particular experiments.
1. Add 100 μ l of Standard, Blank, or Sample per well. Cover with the Plate sealer. Incubate for 2 hours at 37 C .
2. Remove the liquid of each well, don ’ t wash.
3. Add 100 μ l of Detection Reagent A working solution to each well. Cover with the Plate sealer. Incubate for 1 hour at 37 °C. Detection Reagent A working solution may appear cloudy. Warm to room temperature and mix gently until solution appears uniform.
4. Aspirate each well and wash, repeating the process three times for a total of three washes. Wash by filling each well with Wash Buffer (approximately 400 μ l) using a squirt bottle, multi-channel pipette, manifold dispenser 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.
5. Add 100 μ l of Detection Reagent B working solution to each well. Cover with a new Plate sealer. Incubate for 1 hours at 37 C .
6. Repeat the aspiration/wash as in step
4. 7. Add 90 μ l of Substrate Solution to each well. Cover with a new Plate sealer. Incubate within 30 minutes at 37 °C. Protect from light. 4
8. Add 50 μ l of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
9. Determine the optical density of each well at once, using a microplate reader set to 450 nm.
Important Note:1. Absorbance is a function of the incubation time. Therefore, prior to starting the assay it is recommended that all reagents should be freshly prepared prior to use and all required strip-wells are secured in the microtiter frame. This will ensure equal elapsed time for each pipetting step, without interruption.
2. Please carefully reconstitute Standards or working Detection Reagent A and B according to the instruction, and avoid foaming and mix gently until the crystals have completely dissolved. The reconstituted Standards can be used only once. This assay requires pipetting of small volumes. To minimize imprecision caused by pipetting, ensure that pipettors are calibrated. It is recommended to suck more than 10 μ l for once pipetting.
3. To ensure accurate results, proper adhesion of plate sealers during incubation steps is necessary. Do not allow wells to sit uncovered for extended periods between incubation steps. Once reagents have been added to the well strips, DO NOT let the strips DRY at any time during the assay.
4. For each step in the procedure, total dispensing time for addition of reagents to the assay plate should not exceed 10 minutes.
5. To avoid cross-contamination, change pipette tips between additions of each standard level, between sample additions, and between reagent additions. Also, use separate reservoirs for each reagent.
6. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance readings.
7. Duplication of all standards and specimens, although not required, is recommended.
8. Substrate Solution is easily contaminated. Please protect it from light.

Ergebnisberechnung:

5 Average the duplicate readings for each standard, control, and sample and subtract the average zero standard optical density. Create a standard curve by reducing the data using computer software capable of generating a four parameter logistic (4-PL) curve-fit. As an alternative, construct a standard curve by plotting the mean absorbance for each standard on the x-axis against the concentration on the y-axis and draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the GUS concentrations versus the log of the O.D. and the best fit line can be determined by regression analysis. It is recommended to use some related software to do this calculation, such as curve expert 13.0. This procedure will produce an adequate but less precise fit of the data. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Handhabung: 1. The kit should not be used beyond the expiration date on the kit label.
2. Do not mix or substitute reagents with those from other lots or sources.
3. If samples generate values higher than the highest standard, further dilute the samples with the Assay Diluent and repeat the assay. Any variation in standard diluent, operator, pipetting technique, washing technique,incubation time or temperature, and kit age can cause variation in binding.
4. This assay is designed to eliminate interference by soluble receptors, ligands, binding proteins, and other factors present in biological samples. Until all factors have been tested in the Immunoassay, the possibility of interference cannot be excluded.

Lagerung: 4 °C/-20 °C

Informationen zur Lagerung: The Standard, Detection Reagent A, Detection Reagent B and the 96-well strip plate should be stored at -20 °C upon being received. The other reagents can be stored at 4 °C.

Details zu SOCS1

Target: SOCS1 (Suppressor of Cytokine Signaling 1 (SOCS1))

Andere Bezeichnung: SOCS1 (SOCS1 Produkte)

Synonyme: socs1 ELISA Kit, zgc:91868 ELISA Kit, jab ELISA Kit, cis1 ELISA Kit, ssi1 ELISA Kit, tip3 ELISA Kit, cish1 ELISA Kit, ssi-1 ELISA Kit, socs-1 ELISA Kit, SOCS1 ELISA Kit, LOC100228306 ELISA Kit, socs1b ELISA Kit, CIS1 ELISA Kit, CISH1 ELISA Kit, JAB ELISA Kit, SOCS-1 ELISA Kit, SSI-1 ELISA Kit, SSI1 ELISA Kit, TIP3 ELISA Kit, Cish1 ELISA Kit, Cish7 ELISA Kit, Socs-1 ELISA Kit, suppressor of cytokine signaling 1a ELISA Kit, suppressor of cytokine signaling 1 ELISA Kit, suppressor of cytokine signaling 1 L homeolog ELISA Kit, socs1a ELISA Kit, socs1 ELISA Kit, SOCS1 ELISA Kit, socs1.L ELISA Kit, Socs1 ELISA Kit

Hintergrund: Glucuronidases are members of the glycosidase family of enzymes that catalyze breakdown of complex carbohydrates. Human beta -glucuronidase is a type of glucuronidase (a member of glycosidase Family 2) that catalyzes hydrolysis of beta -D-glucuronic acid residues from the non-reducing end of mucopolysaccharides (also referred to as glycosaminoglycans) such as heparan sulfate. Human beta -glucuronidase is located in the lysosome.

Gen-ID: 4166

Pathways: JAK-STAT Signalweg, Interferon-gamma Pathway, TLR Signalweg, Response to Growth Hormone Stimulus