IGFBP5 ELISA Kit

Produktnummer ABIN625131

Produktdetails IGFBP5 ELISA Kit

Target: IGFBP5 (Insulin-Like Growth Factor Binding Protein 5 (IGFBP5))

Reaktivität: Maus

Nachweismethode: Colorimetric

Methodentyp: Sandwich ELISA

Detektionsbereich: 5-3000 pg/mL

Untere Nachweisgrenze: 5 pg/mL

Applikation: ELISA

Verwendungszweck: Mouse IGFBP-5 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Proben: Serum, Plasma, Cell Culture Supernatant

Analytische Methode: Quantitative

Spezifität: This ELISA kit shows no cross-reactivity with the following cytokines tested: Mouse CD30, L CD30, T CD40, CRG-2, CTACK, CXCL16, Eotaxin , Eotaxin-2, Fas Ligand, Fractalkine, GCSF, GM-CFS, IFN- gamma, IGFBP-3, IGFBP-6, IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-3 Rb, IL-4, IL-5, IL-9, IL-10, IL-12 p40/p70, IL-12 p70, IL-13, IL-17, KC, Leptin R, LEPTIN(OB), LIX, L-Selectin, Lymphotactin, MCP-1, MCP-5, M-CSF, MIG, MIP-1 alpha, MIP-1 gamma, MIP-2, MIP-3 beta, MIP-3 alpha, PF-4, P-Selectin, RANTES, SCF, SDF-1 alpha, TARC, TCA-3, TECK, TIMP-1, TNF RI, TNF RII, TPO, VCAM-1, VEGF.

Sensitivität: < 5 pg/mL

Produktmerkmale:

• Strip plates and additional reagents allow for use in multiple experiments
• Quantitative protein detection
• Establishes normal range
• The best products for confirmation of antibody array data

Bestandteile:

• Pre-Coated 96-well Strip Microplate
• Wash Buffer
• Stop Solution
• Assay Diluent(s)
• Lyophilized Standard
• Biotinylated Detection Antibody
• Streptavidin-Conjugated HRP
• TMB One-Step Substrate

Benötigtes Material:

• Distilled or deionized water
• Precision pipettes to deliver 2 μL to 1 μL volumes
• Adjustable 1-25 μL pipettes for reagent preparation
• 100 μL and 1 liter graduated cylinders
• Tubes to prepare standard and sample dilutions
• Absorbent paper
• Microplate reader capable of measuring absorbance at 450nm
• Log-log graph paper or computer and software for ELISA data analysis

Anwendungsinformationen

Applikationshinweise: Recommended Dilution for serum and plasma samples100 - 1,000 fold

Probenmenge: 100 μL

Plattentyp: Pre-coated

Protokoll:

1. Prepare all reagents, samples and standards as instructed in the manual.
2. Add 100 μL of standard or sample to each well.
3. Incubate 2.5 h at RT or O/N at 4 °C.
4. Add 100 μL of prepared biotin antibody to each well.
5. Incubate 1 h at RT.
6. Add 100 μL of prepared Streptavidin solution to each well.
7. Incubate 45 min at RT.
8. Add 100 μL of TMB One-Step Substrate Reagent to each well.
9. Incubate 30 min at RT.
10. Add 50 μL of Stop Solution to each well.
11. Read at 450 nm immediately.

Aufbereitung der Reagenzien:

1. Bring all reagents and samples to room temperature (18 - 25 °C) before use.
   2. Sample dilution: If your samples need to be diluted, Assay Diluent C (Item L) should be used for dilution of serum/plasma/culture supernatants. Suggested dilution for normal serum/plasma: 100-1000 fold. Please note that levels of the target protein may vary between different specimens. Optimal dilution factors for each sample must be determined by the investigator.
   3. Assay Diluent B (Item E) should be diluted 5-fold with deionized or distilled water before use.
   4. Preparation of standard: Briefly spin the vial of Item C. Add 400 µL Assay Diluent C (Item L) into Item C vial to prepare a 100 ng/mL standard solution. Dissolve the powder thoroughly by a gentle mix. Add 30 µL IGFBP-5 standard from the vial of Item C, into a tube with 970 µL Assay Diluent C to prepare a 3,000 pg/mL standard solution. Pipette 400myl Assay Diluent C into each tube. Use the stock standard solution to produce a dilution series . Mix each tube thoroughly before the next transfer. Assay Diluent C serves as the zero standard (0 pg/mL). 200 µL 30 µL standard + 970 µL 200myl 200 µL 200 µL 200 µL 200 µL 3,000 1,000 333.3 111.1 37.04 12.35 4.12 0 pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL
   5. If the Wash Concentrate (20x) (Item B) contains visible crystals, warm to room temperature and mix gently until dissolved. Dilute 20 ml of Wash Buffer Concentrate into deionized or distilled water to yield 400 ml of 1x Wash Buffer.
   6. Briefly spin the Detection Antibody vial (Item F) before use. Add 100 µL of 1x Assay Diluent B (Item E) into the vial to prepare a detection antibody concentrate. Pipette up and down to mix gently (the concentrate can be stored at 4 °C for 5 days). The detection antibody concentrate should be diluted 80-fold with 1x Assay Diluent B and used in step 4 of Part VI Assay Procedure.
   7. Briefly spin the HRP-Streptavidin concentrate vial (Item G) and pipette up and down to mix gently before use. HRP-Streptavidin concentrate should be diluted 200-fold with 1x Assay Diluent B (Item E). For example: Briefly spin the vial (Item G) and pipette up and down to mix gently . Add 50 µL of HRP-Streptavidin concentrate into a tube with 10 ml 1x Assay Diluent B to prepare a 200-fold diluted HRP-Streptavidin solution (don't store the diluted solution for next day use). Mix well.

Testdurchführung:

1. Bring all reagents and samples to room temperature (18 - 25 °C) before use. It is recommended that all standards and samples be run at least in duplicate.
   2. Add 100 µL of each standard (see Reagent Preparation step 2) and sample into appropriate wells. Cover well and incubate for 2.5 hours at room temperature or over night at 4 °C with gentle shaking.
   3. Discard the solution and wash 4 times with 1x Wash Solution. Wash by filling each well with Wash Buffer (300 myl) using a multi-channel Pipette 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.
   4. Add 100 µL of 1x prepared biotinylated antibody (Reagent Preparation step 6) to each well. Incubate for 1 hour at room temperature with gentle shaking.
   5. Discard the solution. Repeat the wash as in step
   6. Add 100 µL of prepared Streptavidin solution (see Reagent Preparation step 7) to each well. Incubate for 45 minutes at room temperature with gentle shaking.
   7. Discard the solution. Repeat the wash as in step
   8. Add 100 µL of TMB One-Step Substrate Reagent (Item H) to each well. Incubate for 30 minutes at room temperature in the dark with gentle shaking.
   9. Add 50 µL of Stop Solution (Item I) to each well. Read at 450 nm immediately.

Ergebnisberechnung:

Calculate the mean absorbance for each set of duplicate standards, controls and samples, and subtract the average zero standard optical density. Plot the standard curve on log-log graph paper or using Sigma plot software, with standard concentration on the x-axis and absorbance on the y-axis. Draw the best-fit straight line through the standard points.
Typical Data: These standard curves are for demonstration only. A standard curve must be run with each assay. Mouse IGFBP-5 concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10 Assay Diluent C
Sensitivity: The minimum detectable dose of IGFBP-5 is typically less than 5 pg/mL.
Recovery: Recovery was determined by spiking various levels of IGFBP-5 into normal Mouse serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 115.3 86-130 Plasma 95.11 83-102 Cell culture media 91.26 80-101
Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 73.54 98.44 97.32 Range ( %) 68-86 87-106 85-104 1:4 Average % of Expected 71.58 130.5 137.6 Range ( %) 65-84 118-140 123-145
Reproducibility: Intra-Assay: CV<10 % Inter-Assay: CV<12 %

Testpräzision: Intra-Assay: CV< 10 % Inter-Assay: CV< 12 %

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Handhabung: Avoid repeated freeze-thaw cycles.

Lagerung: -20 °C

Informationen zur Lagerung: The entire kit may be stored at -20°C for up to 1 year from the date of shipment. Avoid repeated freeze-thaw cycles. The kit may be stored at 4°C for up to 6 months. For extended storage, it is recommended to store at -80°C.

Haltbarkeit: 6 months

Referenzen für IGFBP5 ELISA Kit (ABIN625131)

Talbot, Sparks, Powell, Kahl, Caperna: "Quantitative and semiquantitative immunoassay of growth factors and cytokines in the conditioned medium of STO and CF-1 mouse feeder cells." in: In vitro cellular & developmental biology. Animal, Vol. 48, Issue 1, pp. 1-11, (2012) (PubMed).

Details zu IGFBP5

Target: IGFBP5 (Insulin-Like Growth Factor Binding Protein 5 (IGFBP5))

Andere Bezeichnung: IGFBP-5 (IGFBP5 Produkte)

Synonyme: IBP5 ELISA Kit, AI256729 ELISA Kit, AW208790 ELISA Kit, IGFBP-5 ELISA Kit, IGFBP-5P ELISA Kit, IGF-BP5 ELISA Kit, igfbp5 ELISA Kit, ibp5 ELISA Kit, IBP-5 ELISA Kit, xIGFBP-5 ELISA Kit, IGFBP5 ELISA Kit, igfbp5b ELISA Kit, zgc:165472 ELISA Kit, insulin like growth factor binding protein 5 ELISA Kit, insulin-like growth factor binding protein 5 ELISA Kit, insulin like growth factor binding protein 5 L homeolog ELISA Kit, insulin-like growth factor binding protein 5b ELISA Kit, insulin-like growth factor-binding protein 5 ELISA Kit, insulin-like growth factor binding protein 5a ELISA Kit, IGFBP5 ELISA Kit, Igfbp5 ELISA Kit, igfbp5.L ELISA Kit, igfbp5b ELISA Kit, igfbp5 ELISA Kit, LOC100194500 ELISA Kit, igfbp5a ELISA Kit

Hintergrund: The Mouse IGFBP-5 (Insulin-like growth factor binding protein 5) ELISA (Enzyme-Linked Immunosorbent Assay) kit is an in vitro enzyme-linked immunosorbent assay for the quantitative measurement of Mouse IGFBP-5 in serum, plasma and cell culture supernatants. This assay employs an antibody specific for IGFBP-5 coated on a 96-well plate. Standards and samples are pipetted into the wells and IGFBP-5 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-Mouse IGFBP-5 antibody is added. After washing away unbound biotinylated antibody, HRP-conjugated streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of IGFBP-5 bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm. Reproducibility: Intra-Assay: CV<10% Inter-Assay: CV<12%.

Gen-ID: 16011

UniProt: Q07079

Pathways: WNT Signalweg, Carbohydrate Homeostasis, Myometrial Relaxation and Contraction, Regulation of Carbohydrate Metabolic Process, Autophagie, Smooth Muscle Cell Migration, Growth Factor Binding