NT-proANP ELISA Assay Kit


The Eagle Biosciences proANP (1-98) ELISA Assay KIT(enzyme-linked immunoassay kit) is intended for the quantitative determination of human, mouse, or rat proANP(1-98) in EDTA plasma, heparin plasma, serum, urine or cell culture supernatants. The proANP (1-98) ELISA Assay KIT is for research use only and not to be used in diagnostic procedures.

SKU: BI-20892 Categories: , ,

NT-proANP ELISA Assay Kit

NT-proANP ELISA Assay Kit Developed and Manufactured in Austria by Biomedica

Size: 1×96 wells
Sensitivity: 0.05 nmol/L
Dynamic Range: 0.63 – 10 nmol/L
Incubation Time: 3.5 hours
Sample Type: plasma, urine, serum or cell culture supernatant
Sample Size: 10 µL
Species: Human, Mouse, Rat
Alternative Names: Atrial Natriuretic Peptide, N-Terminal ProANP
For Research Use Only

Controls Included

Assay Background for NT-proANP ELISA Assay Kit

N-terminal proatrial natriuretic peptide (NT-proANP 1-98) is the biologically inactive fragment (98 amino acid) of the ANP prohormone. ANP is translated from its gene as a 151-amino acid precursor, pre-proANP. After removal of the 25-amino acid signal peptide, the tissue form of the hormone, a 126-amino acid proANP (γ-ANP) is generated. γ-ANP is proteolytically cleaved to the biologically active ANP (α-ANP) and to the 98 amino acid NT-proANP peptide (also known as proANP 1-98). The major molecular forms of circulating human ANP is thus the 28 amino-acid peptide (α-ANP) that contains a ring structure with a disulfide bridge and the 98-amino acid NT-proANP peptide, which is easier to detect in the circulation due to its longer half-life. Both peptides α-ANP and NT-proANP circulate in equimolar amounts.
ANP is secreted from the heart in response to atrial stretching or through stimulation by angiotensin II and endothelin. The most important stimulus for the release of the hormone into circulation is stretching of myocyte fibres. On release, the prohormone is split into equimolar amounts of the highly biologically active proANP (99-126), also known as -ANP or ANP, and the N-terminal part proANP (1-98) (also known as NT-proANP) (Nakagawa et al., 2019; Volpe et al., 2015).

NT-proANP is more stable and has a longer half-life (60-120 min) in circulation than ANP which is rapidly cleared from the circulation with a half-life of 3-4 minutes. The natriuretic peptides have a common characteristic biochemical structure that consists of a ring of 17 amino acids and a disulfide bridge between 2 cysteine molecules.

The ANP prohormone undergoes several cleavages to generate the biologically active form of the hormone. N-terminal (NT) prohormone fragments of natriuretic peptides are typically more stable, have longer half-lives, and circulate at higher concentrations compared to C-terminal biologically active hormone ends (Yandle et al., 1986). It has therefore been suggested that measuring NT prohormone fragments of ANP provide more accurate concentrations in samples (Clerico et al., 2011).
The expression and secretion of ANP increase significantly in pathological states accompanied by stretching the heart chambers, volume overload, and ischemic injury, such as heart failure and myocardial infarction.

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Additional Information

Assay Principle

This kit is a sandwich enzyme immunoassay for the quantitative determination of NT-proANP in human serum, EDTA plasma, heparın plasma, cell culture supernatants and urine. The assay has also been validated for rat samples. In a first step, sample and conjugate (sheep anti-human NT-proANP-HRP) are pipetted into the wells of the microtiter strips, which are pre-coated with polyclonal sheep anti-NT-proANP antibody. NT-proANP present in the sample binds to the pre-coated antibody in the well and forms a sandwich with the conjugate. In the washing step all non-specific unbound material is removed. In a second step, the substrate (TMB Tetramethylbenzidine) is pipetted into the wells. The enzyme catalyzed color change of the substrate is directly proportional to the amount of NT-proANP present in the sample. This color change is detectable with a standard microtiter plate ELISA reader.

The NT-proANP ELISA kit uses highly purified, epitope-mapped antibodies. The antibodies utilized in the NT-proANP ELISA (BI-20892) are as follows:

Capture antibody: AA 1-30 (polyclonal sheep anti-human NT-proANP 1-98)
Detection antibody: AA 79-98 (HRP-labeled polyclonal sheep anti-human NT-proANP 1-98)

Assay Procedure

  1. All reagents and samples must be at room temperature (18-26°C) before use in the proANP (1-98) ELISA Assay Kit.
  2. Mark position for BLANK/STD/SAMPLE/CTRL (Blank/Standard/Sample/Control) on the protocol sheet.
  3. Take microtiter strips out of the bag, take a minimum of one well as Blank. Store unused strips with desiccant at 2-8°C in the bag. Strips are stable until expiry date stated on the label.
  4. Add 10 µl STD/SAMPLE/CTRL (Standards/Sample/Control) in duplicate into respective wells, except blank.
  5. Add 200 µl CONJ (Conjugate) into each well except blank, swirl gently.
  6. Cover tightly and incubate for 3 hrs at room temperature (18-26°C) in the dark.
  7. Aspirate and wash wells 5x with 300 µl diluted WASHBUF (Wash buffer), remove remaining WASHBUF by hitting plate against paper towel after the latest wash.
  8. Add 200 µl SUB (Substrate) into each well.
  9. Incubate for 30 min at room temperature (18-26°C) in the dark.
  10. Add 50 µl STOP (Stop solution) into each well.
  11. Measure absorbance immediately at 450 nm with reference 620 nm, if available.

The assay also detects mouse and rat proANP (1-98)

Package Inserts

Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.

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