Lipopolysaccharide (LPS) is one of the main components of Gram-negative bacteria, which is also known as endotoxin. LPS can cause fever, shock, and even lead to death. With the growing popularity of genetic engineering techniques in biological products and reagents, LPS becomes an important indicator to monitor residual bacteria as well as pyrogenicity in bio-products. This LPS BioAssay™ ELISA Kit has been designed using proprietary hybridoma technology and is an simpler alternative to the traditional LAL (Limulus Amebocyte Lysate) agglutination assay. Following is a comparison between our LPS ELISA Kit and LAL assay: LAL AgglutinationLPS ELISAPrincipleLAL reacts with endotoxinSpecific antibody:antigen interactionQuantitationSemi, visualSpecific, microplate readerThroughput188VectorTest tube96 well plateDilutionMultipleSingleCalculationComplexSimpleSample Volume>100ul 50ul
The Lipopolysaccharide (LPS) BioAssay™ ELISA Kit is a competitive inhibition enzyme immunoassay for the in vitro quantitative measurement of LPS in serum, plasma, tissue homogenates, cell lysates, cell culture supernatants and other biological fluids.
Detection Range:12.35-1000ng/ml
Sensitivity:5.41ng/ml
Intra-Assay CV:<10%
Inter-Assay CV:<12%
Test Principle:This assay employs the competitive inhibition enzyme immunoassay technique. A monoclonal antibody specific for LPS has been pre-coated onto a microplate. A competitive inhibition reaction occurs between biotin-labeled LPS and unlabeled LPS (standards or samples) with the pre-coated antibody specific for LPS. After incubation the unbound conjugate is washed off. Next, avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. The amount of bound HRP conjugate is inversely proportional to the concentration of LPS in the sample. After addition of the substrate solution, the intensity of the color developed is inversely proportional to the concentration of LPS in the sample.
Kit Components:*026552A: Microtiter Strips, 1x96 wells *026552B: Standard, 2x1 vial 026552C: Standard Diluent, 1x10ml*026552D: Detection Reagent A (green), 1x120ul*026552E: Detection Reagent B (red), 1x120ul 026552F: Assay Diluent A, 1x12ml026552G: Assay Diluent B, 1x12ml026552H: TMB Substrate, 1x4.5ml*026552J: Positive Control 584ng/ml, 1x1vial (Lyophilized)026552K; Stop Solution, 1x6ml026552L: Wash Buffer, 30X, 1x20ml
Precaution:The Stop Solution (026552K) suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.
Storage and Stability:Store *026552A, *026552B, *026552D and *026552E at -20°C. Store all the other components at 4°C. Unused kit is stable for 6 months after receipt. Once kit components are opened, it is highly recommended to use remaining reagents within 1 month provided this is within the expiration date of the kit. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap.
Notes on 026552J - Positive Control:Intended Use:The control vials are from a human serum sample known to contain the target protein. It will provide a positive result in the ELISA assay, which indicates that the kit is optimized and working properly.
Characteristics:The control vials contain human lipopolysaccharide in human serum (lyophilized powder). Note: The human based serum used in this product are screened for HIV, Hepatitis A, B, C, HTLV and Syphilis.
Expected Value:584ng/ml
Usage:Dissolve with 150ul standard diluent and let stand for 10 minutes. Mix gently before use. In the experiment, add 50ul to an appropriate well of the plate and test its concentration along with standards and samples. Note: Upon receipt store Positive Control vial at -20°C. After reconstitution, store Positive Control vial at 4°C and use within 5 days.
Assay Procedure Summary:1. Prepare all reagents, samples and standards
2. Add 50ul standard or sample to each well, then add 50ul prepared Detection Reagent A immediately. Shake and mix. Incubate 1 hour at 37°C
3. Aspirate and wash 3 times
4. Add 100ul prepared Detection Reagent B. Incubate 30 minutes at 37°C
5. Aspirate and wash 5 times
6. Add 90ul Substrate Solution. Incubate 10-20 minutes at 37°C
7. Add 50ul Stop Solution. Read at 450nm immediately.
