S100B ELISA Kit

Catalog No. ABIN956114

Product Details S100B ELISA Kit

Target: S100B (S100 Calcium Binding Protein B (S100B))

Reactivity: Human

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Purpose: The Human S-100 ELISA is for the quantitative determination of S-100 in human plasma.

Analytical Method: Quantitative

Specificity: The ELISA kit shows 1% cross-reactivity to Human S-100 and 74% to Human S-100 .

Characteristics: This ELISA kit for determination of human S-100 in plasma samples is based on the sandwich enzyme immunoassay. During the first immune reaction, the human S-100 in calibrators or samples binds to the rabbit anti-bovine S-100 antibodies which are coated on the surface of the microtiter plate. After incubation and plate washing, labeled antibody (biotinylated rabbit anti-bovine S-100 antibodies) are added to bind to the antigen-antibody complex. Then HRP-labeled streptoavidin is added to form biotinylated rabbit anti-bovine S-100beta-antigen-antibody complexes. Finally, HRP enzyme activity is determined by o-phenylenediamine dihydrochloride (OPD) and the concentration of human S-100 is calculated.

Components: 1. Antibody-Coated Plate MTP: 1 plate (96-well) Rabbit anti-bovine S-100beta
2. S-100beta Calibrator Lyophilized: 1 vial (6.3 ng) Bovine S-100beta
3. Labeled Antibody Liquid: 1 bottle (11 mL) Biotinylated rabbit anti-bovine S-100beta
4. SA-HRP solution Liquid: 1 bottle (11 mL) HRP labeled streptoavidin
5. Substrate Buffer Liquid: 1 bottle (26 mL) Citrate buffer containing 0.015% hydrogen peroxide
6. OPD Tablet: 2 tablets o-Phenylenediamine dihydrochloride
7. Stop Solution Liquid: 1 bottle (11 mL) 2N H2SO4
8. Buffer Solution Liquid: 1 bottle (30 mL) Phosphate buffer
9. Wash Solution Liquid: 1 bottle (50 mL) Concentrated saline (Concentrate)
10. Plate Seal: 4 sheets

Material not included: Photometer for microtiter plate (plate reader), which can read absorbance up to 2.5 at 492 nm
Microtiter plate shaker
Washing device for microtiter plate, dispenser with aspiration system
Micropipettes, multi-channel pipettes for 8 wells or 12 wells and their tips
Glass test tubes for preparation of Calibrator Solution
Graduated cylinder (1,000 mL)
Distilled water or de-ionized water

Application Details

Plate: Pre-coated

Reagent Preparation:

1. Preparation of Calibrator: Reconstitute the calibrator (lyophilized S-100 6.3ng/vial) with 1 mL of buffer solution, which makes a 6,300 pg/mL calibrator solution. The reconstituted calibrator solution is to be diluted with the same volume of buffer solution (e.g. 0.2 mL calibrator + 0.2 mL buffer solution), to make a 3,150 pg/mL calibrator solution. Repeat the dilution to make each calibrator of 1,575, 788, 394, 197 and 98 pg/mL. Buffer solution is to be used as the 0 pg/mL calibrator. Note: Calibrator Solution must be prepared immediately before assay. Use clean test tubes or vessels.
   2. Preparation of Substrate Solution: Dissolve one OPD Tablet with 12 mL of Substrate Buffer. Note: Substrate Solution must be prepared immediately before assay. Use clean test tubes or vessels.
   3. Preparation of Wash Solution: Dilute 50 mL of Wash Solution Concentrate to 1,000 mL with distilled or de-ionized water. Diluted Wash Solution is stable for 6 months at 4° C. Note: During storage of the Wash Solution Concentrate at 4° C, precipitates may be observed, however, they will dissolve when diluted.
   4. Other reagents are ready for use.

Sample Preparation:

Samples must be used as soon as possible after collection. If the samples are to be tested at a later time, they should be divided into test tubes in small amounts and frozen at or below - 30° C. Avoid repeated freeze/thaw cycles.

Assay Procedure:

1. Bring all reagents and samples to room temperature (20-30° C) before starting the assay.
   2. Discard or aspirate the solution in the wells and wash the wells 3 times with approximately 0.3 mL of wash solution. Finally, invert the plate and tap it onto an absorbent surface, such as paper towels, to ensure removal of most of the residual wash solution.
   3. Fill 70 µL of buffer solution into all wells first and then introduce 30 µL of calibrator solutions (0, 98, 197, 394, 788, 1,575, 3,150, 6,300 pg/mL) or samples into the wells.
   4. Cover the plate with a Plate Seal and incubate it at 4° C overnight for 15~24 hours. (Shaker not needed)
   5. After incubation, bring the plate to room temperature (2030° C) and let stand for 30 to 40 minutes. Take off the Plate Seal, aspirate and wash the wells four times with approximately 0.3 mL/well of wash solution. Finally, invert the plate and tap it onto an absorbent surface, such as paper towels, to ensure removal of most of the residual wash solution.
   6. Pipette 100 µL of Labeled Antibody into the wells.
   7. Cover the plate with a Plate Seal and incubate it for two hours at room temperature. During the incubation, the plate should be shaken with a microtiter plate shaker.
   8. Take off the Plate Seal, aspirate the solution in the wells and wash the wells four times with approximately 0.3 mL/well of washing solution. Finally, invert the plate and tap it onto an absorbent surface, such as paper towels, to ensure removal of most of the residual wash solution.
   9. Pipette 100 µL of SA-HRP solution into the wells.
   10. Cover the plate with a Plate Seal and incubate it for two hours at room temperature. During the incubation, the plate should be shaken with a microtiter plate shaker.
   11. Dissolve one OPD tablet with 12 mL of substrate buffer. Make sure to prepare immediately before use.
   12. Take off the Plate Seal, aspirate the solution in the wells and wash the wells five times with approximately 0.3 mL/well of washing solution. Finally, invert the plate and tap it onto an absorbent surface, such as paper towels, to ensure removal of most of the residual wash solution.
   13. Pipette 100 µL of the substrate solution containing OPD into the wells, cover with a Plate Seal and incubate the plate for 20 minutes at room temperature for the color reaction.
   14. Add 100 µL of stop solution into the wells to stop color reaction.
   15. Read the optical absorbance of the wells at 492 nm. Calculate the mean absorbance values of the calibrator solutions and plot a calibration curve on semi logarithmc paper (abscissa concentration of calibrator, ordinate: absorbance value). Use the average absorbance of each sample to determine the corresponding value by simple interpolation from the calibration curve. Note: Perform all determinations in duplicate.

Calculation of Results:

Calculate mean absorbance values of wells containing the Calibrators and plot a calibration curve on semilogarithmic graph paper (abscissa: concentration of Calibrators, ordinate: absorbance values of Calibrators). When a sample value exceeds 6,300 pg/mL, it needs to be diluted with buffer solution until the value is within the assay range.

Restrictions: For Research Use only

Handling

Storage: 4 °C

Target Details for S100B

Target: S100B (S100 Calcium Binding Protein B (S100B))

Alternative Name: S-100beta (S100B Products)

Synonyms: NEF ELISA Kit, S100 ELISA Kit, S100-B ELISA Kit, S100beta ELISA Kit, S100P ELISA Kit, AI850290 ELISA Kit, Bpb ELISA Kit, si:dkey-110p14.2 ELISA Kit, wu:fq18b10 ELISA Kit, zgc:92776 ELISA Kit, S100 calcium binding protein B ELISA Kit, S100 protein, beta polypeptide, neural ELISA Kit, S100 calcium binding protein, beta (neural) ELISA Kit, S100B ELISA Kit, S100b ELISA Kit, s100b ELISA Kit

Background: Human S-100 has a molecular weight of 21 kDa and consists of two subunits, chain and chain. It is known that the combination of these subunits is different depending on the location in the human body. S-100 is localized in glial cells and schwann cells, S-100 in glial cells and S-100 in striated muscle, heart and kidney. It was reported that the concentration of S-100 in cerebrospinal fluid was a useful marker for the study of the degree of brain damage after head injury, cerebral hemorrhage and ischemic stroke. Recently, another report described that the increase of S-100 in blood correlated to the degree of brain damage after cerebral ischemia, infarction, hemorrhage and severe head injury.

Pathways: Regulation of Muscle Cell Differentiation, Positive Regulation of Immune Effector Process, Toll-Like Receptors Cascades, Regulation of long-term Neuronal Synaptic Plasticity, S100 Proteins