RFP ELISA Kit

Produktnummer ABIN2344801

Produktdetails RFP ELISA Kit

Target: RFP (Red Fluorescent Protein (RFP))

Reaktivität: Discosoma

Nachweismethode: Colorimetric

Methodentyp: Sandwich ELISA

Applikation: ELISA

Proben: Cell Samples, Tissue Lysate

Analytische Methode: Quantitative

Sensitivität: 150 pg/mL

Produktmerkmale: RFP ELISA Kit is an enzyme immunoassay developed for detection and quantitation of RFP or RFP fusion protein in cell or tissue samples. The quantity of RFP or its variants (including TagRFP, TurboRFP, DsRed, tdTomato, mCherry, mKate, mRuby, mBanana, mOrange, mPlum, and mStrawberry) in an unknown sample is determined by comparing its absorbance with that of a known recombinant RFP standard curve. The kit has detection sensitivity limit of 150 pg/mL RFP. The kit also provides an efficient system for rapid quantitation of RFP lentivirus titer for both viral supernatant and purified virus. Each kit provides sufficient reagents to perform up to 96 assays including standard curve and RFP samples.

Bestandteile:

1. Anti-RFP Antibody Coated Plate : One 96-well strip plate (8 x 12).
2. Biotinylated Anti-RFP Antibody (1000X) : One 15 μL vial of biotinylated antibody recognizing sea anemone Discosoma RFP and its variants.
3. Streptavidin-Enzyme Conjugate : One 20 μL vial.
4. Assay Diluent : One 50 mL bottle.
5. 10X Wash Buffer : One 100 mL bottle. 2
6. Substrate Solution : One 12 mL amber bottle.
7. Stop Solution (Part. No. 310808): One 12 mL bottle.

Box 2 (shipped on blue ice packs)

Benötigtes Material:

1. RFP Sample: cell or tissue lysate
2. 10 μL to 1000 μL adjustable single channel micropipettes with disposable tips
3. 50 μL to 300 μL adjustable multichannel micropipette with disposable tips
4. Multichannel micropipette reservoir
5. Microplate reader capable of reading at 450 nm (620 nm as optional reference wave length)

Anwendungsinformationen

Kommentare:

• Detect as little as 150 pg/mL of RFP
• Recognizes TagRFP, TurboRFP, DsRed, mCherry, mKate, mRuby, mBanana, mOrange, mPlum, mStrawberry, and tdTomato
• Simpler and faster than FACS analysis

Plattentyp: Pre-coated

Aufbereitung der Reagenzien:

1X Wash Buffer: Dilute the 10X Wash Buffer Concentrate to 1X with deionized water. Stir to homogeneity. Anti-RFP Antibody and Streptavidin-Enzyme Conjugate: Immediately before use dilute the Biotinylated Anti-RFP antibody 1:1000 and the Streptavidin-Enzyme Conjugate 1:1000 with Assay Diluent. Do not store diluted solutions.

Testdurchführung:

1. Prepare cell or tissue lysates containing RFP or RFP fusion protein. Note: Because the ELISA kit has a linear range of 156 pg/mL to 10000 pg/mL, we recommend using assay diluent to make series of 2-fold dilutions for each unknown sample.
2. Add 100 μL of RFP sample or RFP standard to the Anti-RFP Antibody Coated Plate. Each RFP sample, RFP standard and blank should be assayed in duplicate.
3. Cover with a plate cover and incubate at room temperature for 2 hours on an orbital shaker.
4. Wash microwell strips 5 times with 250 μL 1X Wash Buffer per well with thorough aspiration between each wash. After the last wash, empty wells and tap microwell strips on absorbent pad or paper towel to remove excess 1X Wash Buffer.
5. Add 100 μL of the diluted Biotinylated Anti-RFP antibody to each well. Incubate at room temperature for 1 hour on an orbital shaker.
6. Wash the strip wells 5 times according to step 4 above.
7. Add 100 μL of the diluted Streptavidin-Enzyme Conjugate to each well. Incubate at room temperature for 1 hour on an orbital shaker.
8. Wash the strip wells 5 times according to step 4 above. Proceed immediately to the next step.
9. Warm Substrate Solution to room temperature. Add 100 L of Substrate Solution to each well, including the blank wells. Incubate at room temperature on an orbital shaker. Actual incubation time may vary from 2-20 minutes. Note: Watch plate carefully, if color changes rapidly, the reaction may need to be stopped sooner to prevent saturation.
10. Stop the enzyme reaction by adding 100 μL of Stop Solution into each well, including the blank wells. Results should be read immediately (color will fade over time).
11. Read absorbance of each microwell on a spectrophotometer using 450 nm as the primary wave length. 4

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Lagerung: 4 °C/-80 °C

Informationen zur Lagerung: Upon receipt, aliquot and store recombinant RFP Standard at -80°C and avoid freeze/thaw. Store all other components at 4°C until their expiration dates.

Referenzen für RFP ELISA Kit (ABIN2344801)

Fang, Liu, Zhang, Sakamoto, Taatjes, Jena, Sun, Woods, Bryson, Kowluru, Zhang, Chen: "COPII-Dependent ER Export: A Critical Component of Insulin Biogenesis and ?-Cell ER Homeostasis." in: Molecular endocrinology (Baltimore, Md.), Vol. 29, Issue 8, pp. 1156-69, (2015) (PubMed).

Details zu RFP

Target: RFP (Red Fluorescent Protein (RFP))

Andere Bezeichnung: RFP (RFP Produkte)

Hintergrund: Red fluorescent protein (DsRed) is a spontaneously fluorescent protein isolated from the Indo-Pacific sea coral, Discosoma striata. It absorbs and emits orange-red light and is well suited for multi-color tagging used in FRET. Since the molecular cloning of RFP cDNA and demonstration of RFP as a functional transgene, RFP has become a powerful tool with exciting applications in developmental, cell and molecular biology. RFP fluorescence is not species specific and can be expressed in bacteria, yeast, plant and mammalian cells. RFP can fuse with proteins of interest without interfering significantly with their assembly and function. Based on the structure of the RFP molecule, many RFP variants have been created with much improved fluorescence emission, or shifted excitation or emission spectra that are well suited for fluorescence microscopy and flow cytometry. Although RFP expression can be easily detected under a fluorescence microscope, RFP fluorescence intensity varies from cell to cell because of the heterogeneity nature of RFP expression. In order to quantitate the RFP expression in cells, FACS analysis is usually required, which is both expensive and time consuming.