TUBB Kit ELISA

N° du produit ABIN454567

Détail du produit TUBB Kit ELISA

Antigène: TUBB (Tubulin, beta (TUBB))

Reactivité: Humain

Méthode de détection: Colorimetric

Type de méthode: Sandwich ELISA

Gamme de detection: 1.56-100 ng/mL

Seuil minimal de détection: 1.56 ng/mL

Application: ELISA

Fonction: This immunoassay kit allows for the in vitro quantitative determination of human beta-tubulin concentrations in tissue homogenates and other biological fluids.

Type d'échantillon: Tissue Homogenate

Analytical Method: Quantitative

Specificité: This assay recognizes recombinant and natural human beta-tubulin.

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

Sensibilité: < 1 ng/mL
The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest detectable concentration that could be differentiated from zero.

Attributs du produit: Homo sapiens,Human,Tubulin beta chain,Tubulin beta-5 chain,TUBB,TUBB5,OK/SW-cl.56

Ingrédients: Reagent (Quantity): Assay plate (1), Standard (2), Sample Diluent (1x20ml), Assay Diluent A (1x10ml), Assay DiluentB 1 x 10ml Detection Reagent A (1x120µl), Detection Reagent B (1x120µl), Wash Buffer(25 x concentrate) (1x30ml), Substrate (1x10ml), Stop Solution (1x10ml)

Information d'application

Volume d'échantillon: 100 μL

Plaque: Pre-coated

Protocole: The microtiter plate provided in this kit has been pre-coated with an antibody specific to beta-tubulin. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated polyclonal antibody preparation specific for beta-tubulin and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB (3,3'5, 5' tetramethyl-benzidine) substrate solution is added to each well. Only those wells that contain beta-tubulin, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of beta-tubulin in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Préparation des réactifs:

Bring all reagents to room temperature before use. 3 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 produces a stock solution of 100 ng/mL. Allow the standard to sit for a minimum of 15 minutes with gentle agitation prior to making serial dilutions. The undiluted standard serves as the high standard (100 ng/mL). The Sample Diluent serves as the zero standard (0 ng/mL). Detection Reagent A and B - Dilute to the working concentration using Assay Diluent A and B (1:100), respectively.

Prélèvement de l'échantillon: Tissue homogenates - The preparation of tissue homogenates will vary depending upon tissue type. For this assay, tissue was rinsed with 1X PBS to remove excess blood, homogenized in 20 mL of 1X PBS and stored overnight at ≤ -20 °C. After two freeze-thaw cycles were performed to break the cell membranes, the homogenates were centrifuged for 5 minutes at 5000 x g. Remove the supernate and assay immediately or aliquot and store at ≤ -20 °C. other biological fluids - Remove particulates by centrifugation and assay immediately or aliquot and store samples at -20 °C or -80 °C. Avoid repeated freeze-thaw cycles. Note: Serum, plasma, and cell culture supernatant samples to be used within 7 days may be stored at 2-8C, 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. It is recommended that all samples be assayed in duplicate.

Procédure de l'essai:

Allow all reagents to reach room temperature. All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Arrange and label required number of strips. Prepare all reagents, working standards and samples as directed in the previous sections.
1. Add 100 uL of Standard, Blank, or Sample per well. Cover with the adhesive strip. Incubate for 2 hours at 37 °C.
2. Remove the liquid of each well, don’t wash.
3. Add 100 uL of Detection Reagent A working solution to each well. 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 (350 uL) 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 uL of Detection Reagent B working solution to each well. Cover with a new adhesive strip.Incubate for 1 hours at 37 °C.
6. Repeat the aspiration/wash as in step
4. 7. Add 90 uL of Substrate Solution to each well. Incubate within 30 minutes at 37°C. Protect from light.
8. Add 50 uL 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. 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.
2. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance readings.
3. It is recommended that no more than 32 wells be used for each assay run if manual pipetting is used since pipetting of all standards, specimens and controls should be completed within 5 minutes. A full plate of 96 wells may be used if automated pipetting is available.
4. Duplication of all standards and specimens, although not required, is recommended.
5. When mixing or reconstituting protein solutions, always avoid foaming.
6. 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.
7. To ensure accurate results, proper adhesion of plate sealers during incubation steps is necessary.
8. Do not substitute reagents from one kit lot to another. Use only the reagents supplied by manufacturer.

Calcul des résultats:

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 y-axis against the concentration on the x-axis and draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the beta-tubulin concentrations versus the log of the O.D. and the best fit line can be determined by regression analysis. 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.

Restrictions: For Research Use only

Stockage

Conseil sur la manipulation: 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.

Stock: 4 °C/-20 °C

Stockage commentaire: 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.

Détail du antigène

Antigène: TUBB (Tubulin, beta (TUBB))

Autre désignation: TUBB (TUBB Produits)

Synonymes: M40 Kit ELISA, OK/SW-cl.56 Kit ELISA, TUBB1 Kit ELISA, TUBB5 Kit ELISA, XLOT Kit ELISA, m40 Kit ELISA, ok/sw-cl.56 Kit ELISA, tubb1 Kit ELISA, tubb5 Kit ELISA, TUBB2A Kit ELISA, TUBB2B Kit ELISA, TUBB Kit ELISA, 1t Kit ELISA, B1t Kit ELISA, BETA 56D Kit ELISA, CG9277 Kit ELISA, DTB2 Kit ELISA, Dmbeta1 Kit ELISA, Dmel\\CG9277 Kit ELISA, T Kit ELISA, Tub Kit ELISA, Tubulin Kit ELISA, beta Kit ELISA, beta-Tub Kit ELISA, beta-Tub56D Kit ELISA, beta-tub Kit ELISA, beta-tubulin56D Kit ELISA, beta1 Kit ELISA, beta1-Tubulin Kit ELISA, beta1-tub Kit ELISA, beta1Tub Kit ELISA, beta1t Kit ELISA, beta1tub Kit ELISA, beta56D Kit ELISA, betaTub Kit ELISA, betaTub1 Kit ELISA, beta[[1]] tubulin Kit ELISA, beta[[1]]-tubulin Kit ELISA, betatub(56D) Kit ELISA, 143391_i_at Kit ELISA, 3t Kit ELISA, B3t Kit ELISA, BETA 60D Kit ELISA, CG3401 Kit ELISA, D.m.BETA-60D Kit ELISA, DTB3 Kit ELISA, Dmbeta3 Kit ELISA, Dmel\\CG3401 Kit ELISA, Tub60D Kit ELISA, beta-Tub60D Kit ELISA, beta-Tub6D Kit ELISA, beta3 Kit ELISA, beta3 TU Kit ELISA, beta3-Tub Kit ELISA, beta3-tubulin Kit ELISA, beta3TUB Kit ELISA, beta3Tub Kit ELISA, beta3t Kit ELISA, beta60C Kit ELISA, betaTub3 Kit ELISA, betaTub60C Kit ELISA, beta[[3]] tubulin Kit ELISA, beta[[3]]-Tub Kit ELISA, beta[[3]]-tubulin Kit ELISA, betatub60D Kit ELISA, p50 Kit ELISA, p50/tubulin Kit ELISA, p53 Kit ELISA, p53/tubulin Kit ELISA, 2t Kit ELISA, B2t Kit ELISA, BETA 85D Kit ELISA, BETA2 Kit ELISA, CG9359 Kit ELISA, D.m.BETA-85D Kit ELISA, DTB4 Kit ELISA, Dmbeta2 Kit ELISA, Dmel\\CG9359 Kit ELISA, beta(2)Tu Kit ELISA, beta(2)Tub Kit ELISA, beta-Tub85D Kit ELISA, beta-tub85D Kit ELISA, beta2 Kit ELISA, beta2-tub Kit ELISA, beta2t Kit ELISA, beta2tub Kit ELISA, beta85D Kit ELISA, betaTub2 Kit ELISA, beta[[2]]-tubulin Kit ELISA, ms(3)KK[D] Kit ELISA, 4t Kit ELISA, B4t Kit ELISA, BETA 98B Kit ELISA, CG4869 Kit ELISA, DTB1 Kit ELISA, Dmbeta4 Kit ELISA, Dmel\\CG4869 Kit ELISA, beta-Tub97EF Kit ELISA, beta4 Kit ELISA, beta4t Kit ELISA, beta97F Kit ELISA, betaTub4 Kit ELISA, betaTub98BC Kit ELISA, betaTub98C Kit ELISA, Tub1 Kit ELISA, bmtub1 Kit ELISA, Tub3 Kit ELISA, bmtub3 Kit ELISA, Tub2 Kit ELISA, bmtub2 Kit ELISA, Tub4 Kit ELISA, bmtub4 Kit ELISA, LOC100101153 Kit ELISA, tubulin beta class I Kit ELISA, putative cobalt-zinc-cadmium outer membrane resistance protein Kit ELISA, Tubulin beta chain Kit ELISA, tubulin beta-2A chain Kit ELISA, beta-Tubulin at 56D Kit ELISA, beta-Tubulin at 60D Kit ELISA, beta-Tubulin at 85D Kit ELISA, beta-Tubulin at 97EF Kit ELISA, beta-tubulin Kit ELISA, tubulin beta 6 class V Kit ELISA, beta tubulin Kit ELISA, tubulin beta class I L homeolog Kit ELISA, tubulin, beta 5 class I Kit ELISA, TUBB Kit ELISA, czcC Kit ELISA, tbb-6 Kit ELISA, tubb Kit ELISA, LOC462399 Kit ELISA, betaTub56D Kit ELISA, betaTub60D Kit ELISA, betaTub85D Kit ELISA, betaTub97EF Kit ELISA, Tub1 Kit ELISA, Tub3 Kit ELISA, Tub2 Kit ELISA, Tub4 Kit ELISA, TVAG_525430 Kit ELISA, TVAG_062880 Kit ELISA, TVAG_456920 Kit ELISA, LOC100101153 Kit ELISA, TUBB6 Kit ELISA, Tb927.1.2330 Kit ELISA, Tb927.1.2350 Kit ELISA, Tb927.1.2370 Kit ELISA, Tb927.1.2390 Kit ELISA, tubb.L Kit ELISA, Tubb Kit ELISA, LOC693049 Kit ELISA, LOC443015 Kit ELISA, Tubb5 Kit ELISA

Sujet: A Tubulin is one of several members of a small family of globular proteins. The most common members of the tubulin family are alpha-tubulin and beta-tubulin, the proteins that make up microtubules. Each has a molecular weight of approximately 55 kiloDaltons. Microtubules are assembled from dimers of alpha- and beta-tubulin. These subunits are slightly acidic with an isoelectric point between 5.2 and 5.8. Tubulin was long thought to be specific to eukaryotes. Recently, however, the prokaryotic cell division protein FtsZ was shown to be evolutionarily related to tubulin. To form microtubules, the dimers of alpha- and beta-tubulin bind to GTP and assemble onto the (+) ends of microtubules while in the GTP-bound state. After being incorporated into the microtubule, the bound molecule of GTP will hydrolyse into GDP. Although both subunits bind GTP, only the beta-subunit has GTPase activity, that is, beta-tubulin can hydrolyse GTP to GDP whereas alpha-tubulin cannot. Whether the beta-tubulin member of the tubulin dimer is bound to GTP or GDP influences the stability of the dimer in the microtubule. Dimers bound to GTP tend to assemble into microtubules, while dimers bound to GDP tend to fall apart, thus, this GTP cycle is essential for the dynamic instability of the microtubule. Class III beta-tubulin is a microtubule element expressed exclusively in neurons, and is a popular identifier specific for neurons in nervous tissue. Katanin is a protein complex that severs beta-tubulin, and is necessary for rapid microtubule transport in neurons and in higher plants.

Pathways: Dynamique des Microtubules, M Phase