Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit
|Pack Size:||96wells/kit, with removable strips.|
|Assay Range:||0.005-0.1 mg/L|
|Sample Type:||Serum and Plasma|
Data & Images
The Bosterbio OneStep ELISA kit is a solid phase direct ELISA sandwich kit. Instead of adding samples detection antibody and ABC-HRP separately, The OneStep ELISA kit allows the user to add standards, samples and controls to wells in one step, along with the incubation buffer. After a simple washing step, an enzyme conjugate reagent is added into each well. After the excess enzyme conjugate is washed out, the substrate is added into each well. The enzyme catalyzes the substrate yielding a blue color (Amax = 370nm and 652nm) that changes to yellow (Amax = 450nm) upon addition of a sulfuric or phosphoric acid stop solution. The intensity of color developed is directly proportional to the concentration of target protein in the samples. A standard curve is generated relating color intensity to the concentration of target protein.
|Product Name||Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit|
|Description||Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit, tested with Serum and Plasma. Format: 96wells/kit, with removable strips.|
|Cite This Product||Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit (Boster Biological Technology, Pleasanton CA, USA, Catalog # EK7040)|
*This antibody is predicted to react with the above species based on antigen sequence similarities. Our Boster Guarantee covers the use of this product with the above species.
*Our Boster Guarantee covers the use of this product in the above tested applications.
**For positive and negative control design, consult "Tissue specificity" under Protein Target Info.
|Recommended Detection Systems||
*Blocking peptide can be purchased at $50. Contact us for more information
**Boster also offers various secondary antibodies for Immunoflourescecne and IHC. Take advantage of the buy 1 primary antibody get 1 secondary antibody for free promotion for the entire year 2018!
|Cross Reactivity||There is no detectable cross-reactivity.|
|Pack Size||96wells/kit, with removable strips.|
*Sensitivity, or Lower Limit of Detection (LLD), is the minimum level of target protein the ELISA assay can detect. We measure 20 blank wells and if the O.D. value is 2 standard deviations higher than the blanks' average O.D. the sample can be deemed positive.
|Assay Range||0.005-0.1 mg/L
*This assay range is determined using common samples. For samples with low target protein concentrations, users can adjust the standard curve to extend the lower limit of assay range.
|Sample Type||Serum and Plasma
*The above listed samples are the ones valided with the assay. If you do not see your sample of interest listed, as long as there is enough level of target protein present in the sample, this Picokine™ ELISA kit should detect it.
**For protocol and tips regarding preparing your sample of interest, please check our ELISA sample preparation guide.
|Normal Levels||Some research articles suggesting the natural levels of CRP are listed below:|
|Storage||Store the kit at 2°C to 8°C. Keep microwells sealed in a dry bag with desiccants. The reagents are stable until expiration of the kit. Do not expose reagent to heat, sun, or strong light. Avoid multiple freeze-thaw cycles(Shipped with wet ice.)|
|1. Microwells coated with CRP MAb||12x8x1|
|2. CRP Standard: 6 vials (ready to use)||0.7ml|
|3. CRP Enzyme Conjugate: 1 bottle (ready to use)||12 ml|
|4. TMB Substrate: 1 bottle (ready to use)||12ml|
|5. Stop Solution: 1 bottle (ready to use)||12ml|
|6. Sample Diluent||50 ml|
|7. 20X Wash concentrate: 1 bottle||25ml|
Material Required But Not Provided
1. Distilled or deionized water
2. Precision pipettes
3. Disposable pipette tips. Multichannel pipettes are recommended in the condition of large amount of samples in the detection.
4. ELISA reader capable of reading absorbance at 450nm
5. Absorbance paper or paper towel
6. Graph paper
Protein Target Info (Source: Uniprot.org)
You can check the tissue specificity below for information on selecting positive and negative control.
|Tissue Specificity||Found in plasma.|
Dilution Ratios/Recommended Concentrations
At Boster we strive to provide the best Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit by testing all applications on non-spiked tissues and cell lines to ensure that the affinity of the antibody is enough to react to the endogenouse level of the target protein. Read more about our QC panel here.
|Recommended dilution ratios are listed below:|
|ELISA Target remove|
**Boster provides high sensitivity secondary antibody kits for Western blotting and IHC. For more info see Related Products below.
Human High Sensitivity C-Reactive Protein (CRP) ELISA Kit Images
Click the images to enlarge.
Intra/Inter Assay Precision
WARNINGS AND PRECAUTIONS
1. Potential biohazardous materials: The calibrator and controls contain human source components which have been tested and found non-reactive for hepatitis B surface antigen as well as HIV antibody with FDA licensed reagents. However, there is no test method that can offer complete assurance that HIV, Hepatitis B virus or other infectious agents are absent. These reagents should be handled at the Biosafety Level 2, as recommended in the Centers for Disease Control/National Institutes of Health manual, "Biosafety in Microbiological and Biomedical Laboratories" 1984.
2. This test kit is designed for research use only.
3. Do not pipette by mouth. Do not smoke, eat, or drink in the areas in which specimens or kit reagents are handled.
5. The components in this kit are intended for use as an integral unit. The components of different lots should not be mixed.
6. It is recommended that standards, control and serum samples be run in duplicate.
7. Optimal results will be obtained by strict adherence to this protocol. Accurate and precise pipetting, as well as following the exact time and temperature requirements prescribed are essential. Any deviation from this may yield invalid data.
SPECIMEN COLLECTION AND HANDLING
1. Collect blood specimens and separate the serum immediately.
2. Typically, specimens may be stored refrigerated at (2-8°C) for 5 days. If storage time exceeds 5 days, store frozen at (-20°C) for up to one month.
3. Avoid multiple freeze-thaw cycles.
4. Prior to assay, frozen sera should be completely thawed and mixed well.
5. Do not use grossly lipemic specimens.
1X Wash Buffer: Prepare 1X Wash buffer by adding the contents of the bottle (25 ml, 20X) to 475 ml of distilled or deionized water. Store at room temperature (20-25°C).
Prior to assay, allow reagents to stand at room temperature (20-25°C). Gently mix all reagents before use.
1. Place the desired number of coated strips into the holder
2. Dilute patient samples and controls 1:100 by adding 5 ul of samples to 495 ul of sample Diluent (STANDARDS ARE READY TO USE).
3. Dispense 10ul of standard, diluted samples and controls into the appropriate wells
4. Add 100ul of enzyme conjugate to all wells. Tap the holder to remove air bubbles from the liquid and mix well.
5. Incubate for 60 minutes at room temperature (20-25°C).
6. Remove liquid from all wells. Wash wells three times with 300ul of 1X wash buffer. Blot on absorbent paper towels.
7. Add 100ul of TMB substrate to all wells.
8. Incubate for 15 minutes at room temperature.
9. Add 50ul of stop solution to all wells. Shake the plate gently to mix the solution.
10. Read absorbance on ELISA Reader at 450 nm within 15 minutes after adding the stopping solution.
CALCULATION OF RESULTS
The standard curve is constructed as follows:
1. Check CRP standard value on each standard vial. This value might vary from lot to lot. Make sure you check the value on every kit. See example of the standard attached.
2. To construct the standard curve, plot the absorbance for the IgE standards (vertical axis) against its versus the CRP standard concentrations (horizontal axis) on a linear graph paper. Draw the best curve through the points.
3. Read the absorbance for controls and each unknown sample from the curve. Record the value for each control or unknown sample.
4. The obtained values of the patient samples and control sera should be multiplied by the dilution factor of 100 to obtain CRP results in mg/l.
5. Patient samples with CRP concentrations greater than 10 mg/l should be further diluted 10-fold after the initial 100-fold dilution (total dilution 1:1,000), and the final CRP values should be multiplied by 1,000 to obtain CRP results in mg/l.
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We promise all of our products perform as described in datasheets.
Q: I found bubbles in the wells of my ELISA kit. Is this normal or will it have an impact on the performance of the kit?A: The bubbles in the wells is an uncommon issue, but it would not affect the results of your experiment.
We suggest to wash the plate to remove the bubbles as suggested in the protocol below prior to the assay:
- Wash the plate 3 times with the 1X wash buffer.
- Discard the liquid in the wells into an appropriate waste receptacle. Then, invert the plate on the benchtop onto a paper towel and tap the plate to gently blot any remaining liquid. It is recommended that the wells are not allowed to completely dry at any time.
- Add 300 µl of the 1x wash buffer to each assay well. (For cleaner background incubate for 60 seconds between each wash).
- Repeat steps a-b 2 additional times.
Q: There are precipitates in the sample diluent buffer. What is it?A: The precipitates are fatty substances contained in BSA. It would not affect the performance of the kit. If you want to confirm this, please contact Boster Support.
Q: I left this product out in ambient temperatures for around 5 days, will the kit be stable?A: Our kits should still be stable after being in ambient temperatures for around 5 days. It would not affect the performance of the kit. We suggest you to run the test and contact us if does not work as expected.
Q: What is the number of density of cells recommended for ELISA assay (for cell culture supernants and cell lysates)?A: The number of cells we recommened for cell culture supernants and cell lysates is 10^6 cells/mL.
Q: How many samples can I test on one ELISA plate?A: On each 96 well plate, we recommend running an 8-point standard curve with duplicate wells. With the remaining 80 wells, 40 samples can be tested in duplicate.
Q: The volumes of my samples are small, would it be possible to use 50µl/sample?A: There are two solutions for insufficient sample volume. <br> 1. Dilute the samples with the provided sample diluent buffer into 100ul. <br> 2. If 50ul of sample is added into a well, add 50ul of standard solution.
Q: Will this kit work on tissue samples?A: For tissue homogenates, endogenous biotin should be considered. Endogenous biotin in tissue homogenates might introduce false positive signal. Please run the test as described below to confirm if there is Endogenous biotin in the tissue lysate samples using the ELISA kit. <br> • Add tissue homogenates into the wells and then add ABC and TMB without adding any biotinylated detection antibody to see if any signal will be observed. <br> • If no signal is produced, then you can work on the tissue sample by using the kit.
Q: Is it possible to dilute 10X TBS stock that is 250mM Tris Base/Tris-Hcl, 27 mM KCl, and 1.37M NaCl to be used as the wash buffer?A: You can dilute the 10X TBS stock to 1XTBS and test the pH value of the 1xTBS. <br> The 1XTBS can be used if the pH value falls in the range of 7.2-7.6.
Q: Would your kits work normally with samples from Wistar Rat?A: Our kits would work normally with samples from Wistar Rat.
Q: How can I store the dissolved wash buffer for a longer period of time?A: You can prepare a concentrated wash buffer (25X) which can be stored at 4°C for 1-3 months.
Q: Does the TMB color developing agent contain H2O2?A: Yes, the TMB color developing agent contain H2O2.
Q: Does this ELISA kit contain any product produced in humans or primates? <br> Keywords: component, ingredientA: None of the components in our ELISA kits are produced in humans or primates.
Q: What is the volume of the recombinant protein control? What is its shelf life? <br>Keywords: expiration, storage, temperature, sizeA: The volume of the control is around 100pg. If unopened, the shelf life of the control is the same as the whole kit - "Store at 4˚C for 6 months, at -20˚C for 12 months." The reconstituted control can be stored at -20˚C for 2 days.
Q: Will it be okay to run the experiment if I accidentally used the wrong buffer (e.g. sample diluent buffer) for antibody dilution instead of the antibody diluent buffer? Keywords: dilution, replace, substitute, sample bufferA: Using the sample diluent buffer for antibody dilution instead of the antibody diluent buffer will negatively affect the test result to some extent. The values of both standard and sample might be lower than the normal values.
Q: The protocol says to use neutral PBS and provides a recipe. Could I use neutral PBS made with KCl or KH2PO4 (common constituents for most PBS recipes) instead? Keywords: protocol, alternative buffer, applications, reagent recipeA: Yes, it is ok to use common PBS recipes. We have tried many types of buffers with common constituents, and no significant difference was observed whatsoever.
Q: Why are my O.D. values different than your values on the datasheet? Keyword: troubleshooting, referenceA: We detected the kit in our lab and got our values on the datasheet before delivery, but protein activity will decrease as time goes on, so you may get lower O.D. values than ours. However, it should still be in a reasonable range and the standards can be used to calculate sample values. Good linearity of curve is more important than the actual numerical value.
Q: On the ELISA kit datasheet it says "HRP substrate TMB was used to visualize HRP enzymatic reaction." What is HRP and which part of the kit contains HRP? Keyword: kit components, reagents, protocolA: It means Avidin-Biotin-Peroxidase Complex(AR1103) and you could find it in Kit Components.
Q: Do you offer any ELISA kits that can work with whole blood samples instead of plasma or urine? Keyword: applicationsA: We test serum and plasma routinely, and there is very little difference between serum, plasma and whole blood. The whole blood also contains proteins we need to test. The kit can be used to test whole blood in theory.
Q: Does silicic acid (formula SiOH4) affect the results of the ELISA assay? Keyword: protocol, reagentsA: The acid may affect the binding of antigen to antibody, it is not recommended to use. That being said, it is unlikely to affect the reaction if the solution remains an overall neutral environment.
Q: Is there lot-to-lot variation of the ELISA kits? What are Boster's general services when I have questions about your kits? Keyword: technical support, help, customer serviceA: The NIBSC/WHO 1st International Standard is evaluated, we always test our kits before delivery, and customers can find the test result on the protocol. If the customer needs technical support from us to analyze their data, please contact us and we ask that you provide us the following information: the lot# and production date, and when did the customer detect the kit.
Q: The results of my standards do not look correct, what could be the problem? Keyword: troubleshoot, protocolA: Double check the protocol for plate washing. It should be examined on three aspects: 1) Did you make the washing buffer correctly? 2) How long did you let the buffer stay in wells? 3) What was the volume of the washing buffer added to each well? In addition, pay attention when adding sample to avoid contamination. And we suggest testing the standards again. Values of standards at low concentration are more affected than that at higher concentration, so customer can still get expected values at high concentration even if there is an error. If problems persist, please contact technical support with the catalog and lot number of your product.
Q: What is the well depth of the 96 well plates for the ELISA kits? Keyword: well capacity, product sizeA: The well depth is 300ul, and the max capacity is 350ul. The height of the well is 1.1 cm, and the internal size is 1 cm.
Q: The kit does not include wash solution, what should I do? Keyword: wash bufferA: Our Elisa kits do not come with wash solution, but we have included information about how to make washing buffer on the datasheet. Please refer to the "Material Required But Not Provided" section. We also offer washing buffer for sale separately (Phosphate Buffered Saline Powder SKU: AR0030).
Q: For your ELISA kits, are the capture and detection antibodies polyclonals or monoclonals? Keyword: antibody clonalityA: This information can be found for each kit under the "Properties" section, and you can find the immunogen sequence information in the "Overview" section.
Q: Do your ELISA kits come with sealants or plate covers? Keyword: storage, sequential useA: The kit can be used within a month sequentially if it's opened and stored at 4 degree. There is no need to use sealants, the plate can be packaged with aluminum foil bag, and for other reagents keep bottle tightly closed.
Q: What is the concentration of Sodium Azide in your Elisa kits? Keyword: preservativeA: Our Elisa kits contain 0.02% Sodium Azide. All of the components except TMB colour developing reagent and stop solution contains 0.02% Sodium azide as the preservative.
Q: Are there positive and negative controls available for my ELISA kit? Keyword: positive control, negative controlA: We can provide a recombinant protein as a control. It costs $50 per control and takes 2 weeks to manufacture. We cannot provide a positive or negative control in serum.
Q: Why do I get positive results for my knockout (KO) model when used as a control?A: The knockout (KO) model may contain truncated forms of the target protein which can be detected by the ELISA assay.
Q: How long do I soak my plate in the wash buffer?A: The plate should soak in 0.3mL PBS or TBS wash buffer for at least 1-2 minutes in an automated wash station.
Q: Can I use Tween in my wash buffer?A: While it is not recommended to use Tween in your wash buffer, small amounts (<0.1% concentration) may decrease background due to insufficient blocking.
Q: What plate type do I use to set up the microplate reader?A: Our plates are made with the Corning costar plates similar to the DNA-BIND 96 -well plates.
Q: What should I use for negative control?A: Please contact us for negative control suggestions. You can also check expression databases such as genecards, uniprot etc. Due to logistic reasons, we do not sell serum or lysates that we use internally for positive or negative control.
Q: Where can I find troubleshooting information? What should I do if I have unexpected bands, high background, no signal, weak signalA: You can find Boster's troubleshoot guides under tech support tab. Please contact us for further assistance on troubleshooting your experiment.
Q: What is the normal level of this protein in my sample of interest?A: We have reviewed literature and have gathered this information for most of our ELISA kits. You can find this information on the product page or contact us if you cannot find it. However this information is only suggestive and cannot replace pilot studies in determining the optimal sample dilution ratio.
Q: Is the plate separable? Can I use only part of the kit and save the rest for later? How many samples can I run with one kit?A: Yes the plate is separable if the kit says that there are removable strips. There are 12 strips of 8 wells each, all removable from the plate. The amount of samples you can run depend on a few factors. In the most common ELISA set up, you will use two strips for standards, and 10 strips for samples using duplicates, which let you run up to 40 samples per kit. Contact us if you have questions regarding other situations.
Q: Does the kit contain sample preparation reagents? How do I prepare my samples?A: Since different sample types require different reagents, we do not include them in the ELISA kit. However we do have each reagent mentioned in the file below available at very reasonable prices. Be sure to check them out. For sample preparation protocols please download the file below: https://www.bosterbio.com/media/pdf/Boster_Sample_Preparation_Protocols.pdf
Q: Can this ELISA kit work on brain tissue homogenate, cell culture supernatant, saliva, urine, serum, whole blood or any other sample type?A: In theory the ELISA kit will work for all sample types if the target protein is present at a level that falls within the linear range of the ELISA kit detection range. We guarantee the kit to work on the sample types that we have tested. If you want dilution ratio suggestions on these sample types please contact our technical support. For sample types that we have not tested for, we suggest you run pilot experiments to decide the optimal sample dilution.
Q: Can this ELISA kit react with the pro-form of the target protein? Can this ELISA kit react with an isoform of the protein?A: In general, unless otherwise specified, the ELISA kit is pan-specific, meaning that it will react with all different forms of the target protein if they share the majority of the target protein's sequence. The capture and detection antibodies are reactive to the entire sequence of the standard protein. You can find the sequence information of the standard protein in the "immunogen" section. For more information about the specificity of the kit for your particular experiment, please contact our technical support.
Q: Can this ELISA kit react with human, mouse, rat or other species?A: If the kit is reactive to another commonly used species (human, mouse, and/or rat), we would have listed it as a separate product. If you want to check cross-reactivity to a species that is not included in the 3 species listed above, please contact our technical support for more information. As a rule of thumb, if the sequences are 90%+ identical, there is a high chance of cross-reactivity for your species of interest.