HSP90 ELISA Kit

Produktnummer ABIN6967313

Produktdetails HSP90 ELISA Kit

Target: HSP90 (Heat Shock Protein 90 (HSP90))

Reaktivität: Ratte

Nachweismethode: Colorimetric

Methodentyp: Sandwich ELISA

Detektionsbereich: 125 pg/mL - 8000 pg/mL

Untere Nachweisgrenze: 125 pg/mL

Applikation: ELISA

Verwendungszweck: For quantitative detection of HSP-90 in serum, plasma, tissue homogenates.

Proben: Plasma, Serum, Tissue Homogenate

Analytische Methode: Quantitative

Spezifität: This assay has high sensitivity and excellent specificity for detection of HSP-90. No significant cross-reactivity or interference between HSP-90 and analogues was observed. Note: Limited by current skills and knowledge, it is difficult for us to complete the cross-reactivity detection between HSP-90 and all the analogues, therefore, cross reaction may still exist.

Sensitivität: 75 pg/mL

Bestandteile:

• Pre-coated, ready to use 96-well strip plate
• Plate sealer for 96 wells
• Standard
• Sample/Standard Dilution Buffer
• Biotin-labeled Antibody (Concentrated)
• Antibody Dilution Buffer
• HRP-Streptavidin Conjugate (SABC)
• SABC Dilution Buffer
• TMB Substrate
• Stop Solution
• Wash Buffer (25 x concentrate)
• Instruction manual

Benötigtes Material:

1. Microplate reader (wavelength:450nm)
2. 37 °C incubator
3. Automated plate washer
4. Precision single and multi-channel pipette and disposable tips
5. Clean tubes and Eppendorf tubes
6. Deionized or distilled water

Anwendungsinformationen

Probenmenge: 100 μL

Plattentyp: Pre-coated

Protokoll:

1. Wash plate 2 times before adding Standard, Sample (diluted at least 1/2 with Sample Dilution Buffer) and Control (blank) wells!
2. Add 100 µL standard or sample to each well and incubate for 90 minutes at 37 °C.
3. Aspirate and wash plates 2 times.
4. Add 100 µL Biotin-labeled antibody working solution to each well and incubate for 60 minutes at 37 °C.
5. Aspirate and wash plates 3 times.
6. Add 100 µL SABC Working Solution into each well and incubate for 30 minutes at 37 °C.
7. Aspirate and wash plates 5 times.
8. Add 90 µL TMB Substrate Solution. Incubate 10-20 minutes at 37 °C.
9. Add 50 µL Stop Solution. Read at 450nm immediately and calculation.

Aufbereitung der Reagenzien:

• Bring all reagents and samples to room temperature for 20 minutes before use.
• Wash Buffer: If crystals have formed in the concentrate, you can warm it with 40 °C water bath Concentrated Wash Buffer into 750 mL Wash Buffer with deionized or distilled water. Put unused solution back at 2-8 °C.
• Standards:
  1. Add 1 mL Sample Dilution Buffer into one Standard tube (labeled as zero tube), keep the tube at room temperature for 10 minutes and mix them thoroughly. Note: If the standard tube concentration higher than the range of the kit,please dilute it and labeled as zero tube.
  2. Label 7 EP tubes with 1/2, 1/4, 1/8, 1/16, 1/32, 1/64 and blank respectively. Add 0.3 mLof the Sample Dilution Buffer into each tube. Add 0.3 mLof the above Standard solution (from zero tube) into 1st tube and mix them thoroughly. Transfer 0.3 mL from 1st tube to 2nd tube and mix them thoroughly. Transfer 0.3 mL from 2nd tube to 3rd tube and mix them thoroughly, and so on. Sample Dilution Buffer was used for the blank control. Note: It is best to use Standard Solutions within 2 hours.
• Preparation of Biotin-labeled Antibody Working Solution:
  Prepare it within 1 hour before experiment.
  1. Calculate required total volume of the working solution: 0.1ml/well x quantity of wells. (Allow 0.1-0.2 mLmore than the total volume.)
  2. Dilute the Biotin-detection antibody with Antibody Dilution Buffer at 1:100 and mix them thoroughly. (i.e. Add 1 µL Biotin-labeled antibody into 99 µL Antibody Dilution Buffer.)
• Preparation of HRP-Streptavidin Conjugate (SABC) Working Solution:
  Prepare it within 30 minutes before experiment.
  1. Calculate required total volume of the working solution: 0.1ml/well x quantity of wells. (Allow 0.1-0.2 mLmore than the total volume.)
  2. Dilute the SABC with SABC Dilution Buffer at 1:100 and mix them thoroughly. (i.e. Add 1 µL of SABC into 99 µL of SABC Dilution Buffer.)

Aufbereitung der Proben:

• It is recommended to use fresh samples without long storage, otherwise protein degradation and denaturationmay occur in these samples, leading to false results. Samples should therefore be stored for a short periodat 2 - 8 °C or aliquoted at -20 °C (≤1 month) or -80 °C (≤ 3 months). Repeated freeze-thawcycles should be avoided. Prior to assay, the frozen samples should be slowly thawed and centrifuged toremove precipitates.
• If the sample type is not specified in the instructions, a preliminary test is necessary to determinecompatibility with the kit.
• If a lysis buffer is used to prepare tissue homogenates or cell culture supernatant, there is a possibilityof causing a deviation due to the introduced chemical substance.The recommended dilution factor is for reference only.
• Please estimate the concentration of the samples before performing the test. If the values are not in therange of the standard curve, the optimal sample dilution for the particular experiment has to be determined.Samples should then be diluted with PBS (pH =7.0-7.2).

Note:

The user should estimate the concentration of target protein in the test sample, and select a proper dilution factor to make the diluted target protein concentration fall in the optimal detection range of the kit. Dilute the sample with the provided dilution buffer, and several trials may be necessary. The test sample must be well mixed with the dilution buffer. And also standard curves and sample should be making in pre-experiment. If samples with very high concentrations, dilute samples with PBS first and then dilute the samples with Sample Dilution. The matrix components in the sample will affect the test results, which it need to be diluted at least 1/2 with Sample Dilution Buffer before testing!

Testdurchführung:

Washing

Manual: Discard the solution in the plate without touching the side walls. Clap the plate on absorbent filter papers or other absorbent material. Fill each well completely with 350 µL wash buffer and soak for 1 to 2 minutes, then aspirate contents from the plate, and clap the plate on absorbent filter papers or other absorbent material.
Automatic: Aspirate all wells, and then wash plate with 350 µL wash buffer. After the final wash, invert plate, and clap the plate on absorbent filter papers or other absorbent material. It is recommended that the washer shall be set for soaking 1 minute. (Note: set the height of the needles, be sure the fluid can be sipped up completely)

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 100 µL of zero tube, 1sttube, 2ndtube, 3rdtube, 4thtube, 5thtube, 6thtube and Sample Dilution Buffer (blank) into the standard wells.
3. Add Samples: Add 100 µL 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 100 µL 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 100 µL 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 90 µL 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 50 µL 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.

Regarding calculation, (the relative O.D.450) = (the O.D.450 of each well) - (the O.D.450 of blank well). The standard curve can be plotted as the relative O.D.450 of each standard solution (Y) vs. the respective concentration of the standard solution (X). The target concentration of the samples can be interpolated from the standard curve. It is recommended to use some professional software to do this calculation, such as Curve Expert 1.3 or 1.4.

Note: If the samples measured were diluted, multiply the dilution factor to the concentrations from interpolation to obtain the concentration before dilution.

Testpräzision: Intra-Assay: CV<8%
Inter-Assay: CV<10%

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Lagerung: 4 °C,-20 °C

Informationen zur Lagerung:

1. For unopened kit: All the reagents should be kept according to the labels on vials. The Reference Standard and the 96-well stripe plate should be stored at -20 °C upon receipt while the other reagents should be stored at 4 °C.
2. For used kit: When the kit is used, the remaining reagents need to be stored according to the above storage condition. Besides, please return the unused wells to the foil pouch containing the desiccant pack, and zip-seal the foil pouch.

Haltbarkeit: 6 months

Details zu HSP90

Target: HSP90 (Heat Shock Protein 90 (HSP90))

Andere Bezeichnung: Heat Shock Protein 90 (HSP90 Produkte)

Synonyme: EL52 ELISA Kit, HSP86 ELISA Kit, HSP89A ELISA Kit, HSP90A ELISA Kit, HSP90N ELISA Kit, HSPC1 ELISA Kit, HSPCA ELISA Kit, HSPCAL1 ELISA Kit, HSPCAL4 ELISA Kit, HSPN ELISA Kit, Hsp89 ELISA Kit, Hsp90 ELISA Kit, LAP2 ELISA Kit, git10 ELISA Kit, swo1 ELISA Kit, HSP90 ELISA Kit, htpG ELISA Kit, SCBAC25F8.08 ELISA Kit, 23.m06066 ELISA Kit, 17.m07646 ELISA Kit, HSP90-1 ELISA Kit, 143198_at ELISA Kit, 83 ELISA Kit, 83K HSP ELISA Kit, DMHSP82 ELISA Kit, E(sev)3A ELISA Kit, E(sina)2 ELISA Kit, HSP82 ELISA Kit, HSP83 ELISA Kit, ORF1 ELISA Kit, Su(Raf)3A ELISA Kit, anon-EST:Liang-2.53 ELISA Kit, anon-WO0068693 ELISA Kit, anon-WO0140519.209 ELISA Kit, clone 2.53 ELISA Kit, en(lz)3C/4C ELISA Kit, hsp84 ELISA Kit, l(3)j5C2 ELISA Kit, ms(3)08445 ELISA Kit, stc ELISA Kit, DmelCG1242 ELISA Kit, CG1242 ELISA Kit, Hsp86 ELISA Kit, Hspca ELISA Kit, 86kDa ELISA Kit, 89kDa ELISA Kit, AL024080 ELISA Kit, AL024147 ELISA Kit, Hsp86-1 ELISA Kit, hsp4 ELISA Kit, hsp86 ELISA Kit, hsp89 ELISA Kit, hsp90 ELISA Kit, hsp90a ELISA Kit, hspc1 ELISA Kit, hspca ELISA Kit, hspn ELISA Kit, lap2 ELISA Kit, Hsp90alpha ELISA Kit, heat shock protein 90 alpha family class A member 1 ELISA Kit, heat shock protein Hsp90 ELISA Kit, Hsp90 chaperone ELISA Kit, Heat shock protein 90 ELISA Kit, heat shock protein 90 ELISA Kit, chaperone protein HtpG ELISA Kit, molecular chaperone HtpG ELISA Kit, Heat Shock Protein 90 ELISA Kit, uncharacterized LOC100384473 ELISA Kit, Heat shock protein 83 ELISA Kit, heat shock protein 90, alpha (cytosolic), class A member 1 ELISA Kit, heat shock protein 90 alpha family class B member 1 ELISA Kit, heat shock protein 90kDa alpha family class A member 1 L homeolog ELISA Kit, heat shock protein HSP 90-alpha ELISA Kit, HSP90AA1 ELISA Kit, hsp90 ELISA Kit, HSP90 ELISA Kit, daf-21 ELISA Kit, htpG ELISA Kit, SCO7516 ELISA Kit, MAP_RS10510 ELISA Kit, TP04_0646 ELISA Kit, TP01_0934 ELISA Kit, APH_RS03525 ELISA Kit, GbCGDNIH1_0315 ELISA Kit, MAV_2118 ELISA Kit, HSP90C ELISA Kit, BBOV_IV008400 ELISA Kit, BBOV_III007380 ELISA Kit, ACICU_00312 ELISA Kit, ECL_01244 ELISA Kit, YE105_C1172 ELISA Kit, pco153543(105) ELISA Kit, Hsp83 ELISA Kit, Hsp90aa1 ELISA Kit, HSP90AB1 ELISA Kit, hsp90aa1.1.L ELISA Kit, LOC108698781 ELISA Kit

Hintergrund: HSP-90, Heat Shock Protein 90, HSP90AA1, Hsp90, HSP86, HSP89A, HSP90A, HSP90N, HSPC1, HSPCA, HSPCAL1, HSPCAL4, HSPN, LAP2, FLJ31884, Heat shock 86 kDa, heat shock 90kD protein 1, alpha, heat shock 90kD protein 1, alpha-like 4, heat shock 90kD protein, alpha-like 4, heat shock protein HSP 90-alpha, Hsp89

UniProt: P82995

Pathways: M Phase, Regulation of Cell Size, Signaling Events mediated by VEGFR1 and VEGFR2, VEGFR1 Specific Signals