Name
LL-37, Human, ELISA kit
Catalog nr
HK321-01
(lot number and expiry date are indicated on the label)
Short description
Cathelicidins are a family of antimicrobial proteins predominantly found in the peroxidase-negative granules of neutrophils. The cathelicidins are synthesized as preproproteins. Within the neutrophils, they are stored in granules as inactive proforms after removal of the signal peptide. The active biologic domains of the cathelicidins generally reside in the C-terminus. The C-terminal antimic...
Size
1 x 96 det.
Application
Manual
Description
Cathelicidins are a family of antimicrobial proteins predominantly found in the peroxidase-negative granules of neutrophils. The cathelicidins are synthesized as preproproteins. Within the neutrophils, they are stored in granules as inactive proforms after removal of the signal peptide. The active biologic domains of the cathelicidins generally reside in the C-terminus. The C-terminal antimicrobial peptides are activated when cleaved from the proforms of the cathelicidins by serine proteases from azurophil granules. Human cationic antimicrobial protein (hCAP)18 is the only human cathelicidin identified to date. The antibacterial C-terminus of hCAP-18, LL37 (37 amino acids), has been shown to exert broad antimicrobial activity towards gram-negative as well as gram-positive bacteria and to have synergistic antibacterial effects with the defensins. For instance, deficiency in saliva LL37 accords with occurrence of periodontal disease in patients with morbus Kostmann. Moreover, it functions as a chemotactic agent for neutrophils, monocytes and T cells. LL-37 is markedly resistant to proteolytic degradation and to a limited extent also cytotoxic towards mammalian cells. LL-37 was demonstrated to be present in plasma in levels from 1.2 - 1.8 µg/ml and to be enhanced in infectious diseases.
Application
The human LL-37 ELISA has been developed for the quantitative measurement of natural and recombinant LL-37 in plasma and cell culture medium. In plasma samples, human LL-37 can be measured accurately if plasma samples are diluted at least 20 times. Most reliable results are obtained if EDTA plasma is used. Please be aware that human LL-37 is released from neutrophils into serum in the process of blood coagulation. This will lead to false positive and difficult to interpret results of serum samples. Therefore it is advised to use 'careful plasma'.
Features
- Minimum concentration which can be measured is 0.14 ng/ml human LL-37.
- Measurable concentration range of 0.14-100 ng/ml.
- Working volume of 100 µl/well.
Typical standard curve
Principle
- The human LL-37 ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 3½ hours.
- The efficient format of a plate with twelve disposable 8-well strips allows free choice of batch size for the assay.
- Samples and standards are captured by a solid bound specific antibody.
- Biotinylated tracer antibody will bind to captured human LL-37.
- Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody.
- Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB).
- The enzyme reaction is stopped by the addition of citric acid.
- The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the human LL-37 standards (log).
- The human LL-37 concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.
Storage and stability
Product should be stored at 4°C. Under recommended storage conditions, product is stable for at least six months. After reconstitution the reagents are stable for 1 month if stored at 2-8°C, except for the standard. After reconstitution the standard is stable for 12 hours. For longer stability we recommend to store aliquots at -20°C.
Recovery
Normal human blood samples (plasma), were spiked with human LL-37 in concentrations of 35 and 0.7 ng/ml. Samples with and without human LL-37 were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for human LL-37 ranged between 79% and 100% (mean 90%).
Linearity
The linearity of the assay was determined by serially diluting a sample containing 35 ng/ml human LL-37. The diluted samples were measured in the assay. The line obtained a slope of 1.185 and a correlation coefficient of 0.999. 
References
1. Martineau, A et al; IFN-γ- and TNF-independent vitamin D-inducible human suppression of mycobacteria: the role of cathelicidin LL-37, J Immunol 2007, 178: 7190
Precautions
For research use only. Not for use in or on humans, animals or for diagnostics. It is the responsibility of the user to comply with all local/state and Federal rules in the use of this product. Hycult Biotech is not responsible for any patent infringements that might result with the use of or derivation of this product.
Also available
References
1. Martineau, A et al; IFN-γ- and TNF-independent vitamin D-inducible human suppression of mycobacteria: the role of cathelicidin LL-37, J Immunol 2007, 178: 7190
Scientific info
LL-37 propagates psoriasis by forming complexes with host DNA
Psoriasis is marked by chronic inflammation and excessive proliferation and turnover of the keratinocyte skin cells. The characteristic epidermal infiltration of neutrophils results from the local production of potent antimicrobial agents, including cathelicidin LL-37. LL-37 is involved in antimicrobial responses and wound-healing. Moreover, LL-37 forms complexes with self-DNA that is released from dying skin cells. In normal skin, these DNA/LL-37 complexes probably remain undetected and without consequenses. But in the presence of infiltrating plasmacytoid dendritic cells (pDC) in psoriatic lesions, these DNA/LL-37 complexes trigger strong interferon-alpha production. This uncovers an important role of an endogenous antimicrobial peptide in breaking innate tolerance to self-DNA. References:
1) Lande R, et al; Plasmacytoid dendritic cells sense self-DNA coupled with antimicrobial peptide. Nature 2007, 449: 564
2) Baumgard N, et al; Skin deep but complex. Nature 2007, 449: 1
Psoriasis is marked by chronic inflammation and excessive proliferation and turnover of the keratinocyte skin cells. The characteristic epidermal infiltration of neutrophils results from the local production of potent antimicrobial agents, including cathelicidin LL-37. LL-37 is involved in antimicrobial responses and wound-healing. Moreover, LL-37 forms complexes with self-DNA that is released from dying skin cells. In normal skin, these DNA/LL-37 complexes probably remain undetected and without consequenses. But in the presence of infiltrating plasmacytoid dendritic cells (pDC) in psoriatic lesions, these DNA/LL-37 complexes trigger strong interferon-alpha production. This uncovers an important role of an endogenous antimicrobial peptide in breaking innate tolerance to self-DNA. References:
1) Lande R, et al; Plasmacytoid dendritic cells sense self-DNA coupled with antimicrobial peptide. Nature 2007, 449: 564
2) Baumgard N, et al; Skin deep but complex. Nature 2007, 449: 1
MSDS
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