Human Mannose Binding Lectin / Mannose Binding Protein ELISA Kit
- SKU:
- HUFI00327
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P11226
- Sensitivity:
- 0.094ng/ml
- Range:
- 0.156-10ng/ml
- ELISA Type:
- Sandwich
- Synonyms:
- MBL2, MBL, MBP, MBP1, MBPD, MBL2D, MBP-C, COLEC1, HSMBPC, L-MBP, MBL-C, L-MBP, mannose-binding protein C, mannose-binding lectin, protein C 2
- Reactivity:
- Human
Description
Human Mannose Binding Lectin / Mannose Binding Protein ELISA Kit
Mannose Binding Lectin (MBL) is a crucial component of the innate immune system, playing a vital role in the recognition and elimination of pathogens. MBL is a soluble pattern recognition molecule that binds to specific carbohydrate structures on the surface of various microorganisms, including bacteria, viruses, fungi, and parasites. The Assay Genie Human Mannose Binding Lectin / Mannose Binding Protein ELISA Kit is a highly sensitive assay for the quantitative measurement of MBL in serum, plasma, cell and tissue lysates.
Key Features
| Save Time | Pre-coated 96 well plate | |
| Quick Start | Kit includes all necessary reagents | |
| Publication Ready | Reproducible and reliable results |
Overview
|
Product Name: |
Human Mannose Binding Lectin / Mannose Binding Protein ELISA Kit |
|
Product Code: |
HUFI00327 |
|
Size: |
96 Assays |
|
Alias: |
MBL2, MBL, MBP, MBP1, MBPD, MBL2D, MBP-C, COLEC1, HSMBPC, L-MBP, MBL-C, L-MBP, mannose-binding protein C, mannose-binding lectin, protein C 2 |
|
Detection Method: |
Sandwich ELISA, Double Antibody |
|
Application: |
This immunoassay kit allows for the in vitro quantitative determination of Human MBP/MBL concentrations in serum plasma and other biological fluids. |
|
Sensitivity: |
0.094ng/ml |
|
Range: |
0.156-10ng/ml |
|
Storage: |
4°C for 6 months |
|
Note: |
For Research Use Only |
Additional Information
|
Recovery |
Matrices listed below were spiked with certain level of Human MBP/MBL and the recovery rates were calculated by comparing the measured value to the expected amount of Human MBP/MBL in samples.
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Linearity |
The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human MBP/MBL and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
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CV(%) |
Intra-Assay: CV<8% |
Kit Components
| Component | Quantity | Storage |
|
ELISA Microplate (Dismountable) |
8x12 strips |
4°C for 6 months |
|
Lyophilized Standard |
2 |
4°C/ -20°C |
|
Sample/Standard Dlution Buffer |
20ml |
4°C |
|
Biotin-labeled Antibody (Concentrated) |
120ul |
4°C (Protection from light) |
|
Antibody Dilution Buffer |
10ml |
4°C |
|
HRP-Streptavidin Conjugate (SABC) |
120ul |
4°C (Protect from light) |
|
SABC Dilution Buffer |
10ml |
4°C |
|
TMB Substrate |
10ml |
4°C (Protection from light) |
|
Stop Solution |
10ml |
4°C |
|
Wash Buffer (25X) |
30ml |
4°C |
|
Plate Sealer |
5 |
- |
Other materials required:
- Microplate reader with 450 nm wavelength filter
- Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
- Incubator
- Deionized or distilled water
- Absorbent paper
- Buffer resevoir
Protein Information
Protocol
*Note: Protocols are specific to each batch/lot. For the exact instructions please follow the protocol included in your kit.
Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.
| Step | Procedure |
|
1. |
Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells! |
|
2. |
Aliquot 0.1ml standard solutions into the standard wells. |
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3. |
Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well. |
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4. |
Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells. |
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5. |
Seal the plate with a cover and incubate at 37 °C for 90 min. |
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6. |
Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2. |
|
7. |
Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall. |
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8. |
Seal the plate with a cover and incubate at 37°C for 60 min. |
|
9. |
Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash. |
|
10. |
Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min. |
|
11. |
Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min. |
|
12. |
Add 90 µl of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color. |
|
13. |
Add 50 µl of Stop solution into each well and mix thoroughly. The color changes into yellow immediately. |
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14. |
Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution. |
Sample Type
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
| Sample Type | Protocol |
|
Serum |
If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. |
|
Plasma |
Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit. |
|
Urine & Cerebrospinal Fluid |
Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid. |
|
Cell culture supernatant |
Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately. |
|
Cell lysates |
Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
|
Tissue homogenates |
The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C. |
|
Tissue lysates |
Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C |
|
Breast Milk |
Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles. |
Mannose BInding Lectin Background
Mannose Binding Lectin
Mannose Binding Lectin (MBL) is a vital component of the innate immune system, serving as a pattern recognition molecule that plays a crucial role in identifying and neutralizing invading pathogens. It is a soluble protein that binds to specific carbohydrate structures on the surface of various microorganisms, including bacteria, viruses, fungi, and parasites. Upon binding, MBL initiates a cascade of immune responses, including activation of the complement system and enhancement of phagocytosis, leading to the elimination of the targeted pathogens. MBL also functions as a bridge between innate and adaptive immunity, modulating the overall immune response. Its significance lies in its ability to provide early defense against a wide range of infections and contribute to the maintenance of immune homeostasis. Studying MBL can offer valuable insights into the mechanisms of host-pathogen interactions and aid in the diagnosis and management of infectious and autoimmune diseases.
Mannose Binding Lectin Function
Research conducted over the past decade has provided significant insights into the multifaceted role of Mannose Binding Lectin (MBL) within the innate immune system. Numerous studies have demonstrated that MBL serves as a critical protein with diverse functions, including the activation of complement, facilitation of opsonophagocytosis (independent of complement), modulation of inflammation, and promotion of apoptosis. MBL exerts its effects through various receptor-mediated effector functions, including complement system activation, direct opsonophagocytosis, MBL-dependent cell-mediated cytotoxicity, apoptosis induction, release of pro-inflammatory cytokines, and production of reactive oxygen species. By recognizing and binding to sugar moieties present on the surface of bacteria, viruses, fungi, and parasites, MBL promotes agglutination of these microorganisms, facilitating their phagocytic clearance. Additionally, MBL triggers the activation of the lectin-complement pathway through MBL-associated proteases. These findings shed light on the intricate mechanisms by which MBL contributes to host defense against pathogens and immune regulation.
Mannose Binding Lectin Pathway
MBL-mediated complement activation constitutes a distinct pathway separate from the classical and alternative pathways. The family of MBL-associated serine proteases (MASPs), including MASP-1, MASP-2, MASP-3, and the truncated form MAp19, have been found to associate with MBL. Among them, MASP-2 has been identified as the primary player in complement activation. When MBL-MASP-2 complexes bind to microbial surfaces that display specific sugar arrays, they become activated. MASP-2 exhibits enzymatic activity similar to C1 esterase, leading to the sequential cleavage of C4 and C2. The resulting C4b fragments covalently bind to the microbial surface and interact with cleaved C2 components, facilitated by MASP-2. This interaction gives rise to the formation of the C4b2a complex, which possesses C3 convertase activity, similar to the classical and alternative pathways. Consequently, it generates opsonic C3b fragments and engages the amplification loop of the complement system. This functional activity of MBL explains why MBL deficiency was initially observed in children with recurrent infections associated with impaired opsonization.
Mannose Binding Lectin ELISA Kit FAQs
What is the Human Mannose Binding Lectin ELISA Kit used for?
The Human Mannose Binding Lectin ELISA Kit is designed for the quantitative measurement of Mannose Binding Lectin (MBL) levels in human biological samples. MBL is an important component of the innate immune system and plays a significant role in host defense against various pathogens. This kit enables researchers and clinicians to assess MBL levels, aiding in the study of immune responses, infectious diseases, and genetic variations related to MBL.
What are the advantages of using Human Mannose Binding Lectin ELISA Kit?
The Mannose Binding Lectin ELISA Kit offers several advantages for accurate and precise MBL measurements. It provides high sensitivity and specificity, ensuring reliable results. The kit is user-friendly, offering a streamlined protocol for easy handling and efficient analysis.
What sample types are compatible with Human Mannose Binding Lectin ELISA kit?
The Human Mannose Binding Lectin ELISA Kit is compatible with various sample types, including serum, plasma, cell lysates, and tissue homogenates. It provides flexibility in sample selection, allowing researchers to analyze MBL levels in different biological matrices.
What are the storage requirements for the Human Mannose Binding Lectin ELISA Kit?
The Human Mannose Binding Lectin ELISA Kit components should be stored according to the instructions provided in the kit manual. Generally, it is recommended to store the kit components at the recommended temperature to ensure their stability and optimal performance.
What should I do if my assay results are not optimal?
If you encounter any issues or have suboptimal assay results, we recommend contacting our dedicated support team for assistance. They will be available to provide troubleshooting guidance, answer your questions, and ensure you achieve the best possible results with the MBL ELISA Kit.
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