Understanding NT-proBNP in Veterinary Diseases: A Comprehensive Guide

What Is NT-proBNP? Decoding the Heart Health Biomarker

NT-proBNP (N-terminal pro b-type natriuretic peptide) is a biomarker used in veterinary medicine to assess heart health in animals. Originating from the heart's ventricles, NT-proBNP is released into the bloodstream in response to changes in heart pressure. Elevated levels of NT-proBNP can indicate heart disease, such as cardiomyopathy or congestive heart failure, especially in cats and dogs. This marker is valuable for diagnosing, monitoring the progression of heart disease, and guiding treatment decisions in animals. Testing for NT-proBNP is non-invasive, often requiring only a blood sample, making it an essential tool in veterinary cardiology for early detection and management of heart conditions.

Diseases Detected by NT-proBNP: Insights into Veterinary Cardiology

This table outlines how NT-proBNP levels correlate with heart disease across various animals, detailing normal ranges, disease implications, and measurement methods.

NT-proBNP Biomarker and Its Implications in Veterinary Diseases

Animals	Normal Range	Possible Diseases Listed (Detail)	Specimens/Biofluids	Measurement Method
Cat	<100 pmol/L	HCM (Hypertrophic Cardiomyopathy);RCM (Restrictive Cardiomyopathy); DCM (Dilated Cardiomyopathy) ; Congestive Heart Failure;Arterial Thromboembolism; Systemic Hypertension	Blood Plasma/Serum	ELISA, Chemiluminescence
Dog	<900 pmol/L	DCM; Valvular Heart Disease;Congestive Heart Failure;Chronic Valvular Disease;Pulmonary Hypertension;Cardiac Arrhythmias	Blood Plasma/Serum	ELISA, Chemiluminescence
Horse	-	Atrial Fibrillation; Equine Metabolic Syndrome-related cardiac changes;Congestive Heart Failure	Blood Plasma/Serum	ELISA, Chemiluminescence
Cattle	-	Limited data available, but potential use in detecting heart failure or stress due to metabolic syndromes in dairy cows	Blood Plasma/Serum	ELISA, Chemiluminescence

Note:

"HCM" stands for Hypertrophic Cardiomyopathy, a common heart disease in cats characterized by thickened heart walls."DCM" is Dilated Cardiomyopathy, observed in dogs, characterized by an enlarged heart with poor contraction.The "Normal Range" values provided are generalized; specific values can vary based on the testing method, equipment used, and individual patient factors. It's important to consult veterinary diagnostic reference materials or laboratories for precise normal ranges.In cases where data is not available ("-"), it indicates that the use of NT-proBNP for those specific animals or diseases is either not common or not well-documented in available literature.

Canine quantitative immunoassay reagent for NT-proBNPF

GeneMedi has created many monoclonal antibodies against canine NT-proBNP. All antibodies were examined as capture and detection antibodies in a sandwich immunoassay to find the best antibody pairing. 96-well plates are coated with capture antibodies, and europium chelates mark detection antibodies. To assess antibody pairings, two types of canine NT-proBNP were utilized as antigens: natural canine NT-proBNP from dog plasma and recombinant canine NT-proBNP (Cat. # GMP-CAN-NT-proBNP-Ag01). High sensitivity was demonstrated by many pairings in identifying native and recombinant NT-proBNP. Table 1 displays the best monoclonal antibody pairs.

 

Capture antibody	Detect antibodies
GMP-CAN-NT-proBNP-Ab01	GMP-CAN-NT-proBNP-Ab02
GMP-CAN-NT-proBNP-Ab05	GMP-CAN-NT-proBNP-Ab02
GMP-CAN-NT-proBNP-Ab01	GMP-CAN-NT-proBNP-Ab06

Table 1. Most sensitive antibody pairs

 

These immunoassays have a sensitivity of 22 pg/mL for measuring recombinant NT-proBNP. Figures 1 and 2 display the suggested pairing calibration curves.  Conversion of 1NT-proBNP concentration: 1 pmol/L = 1÷10.545 pg/ml Please note that a variety of factors might affect how well an immunoassay test performs. These consist of the labeling procedure, the kind of label coupled to the detection antibody, and the detection platform. Thus, on our customers' immunoassay systems, other NT-proBNP antibody pairs with non-overlapping epitopes could perform better.

Figure 1, 2. NT-proBNP immunoassay calibration curve

(A) Calibration curve for optimal paired detection

(B) Calibration curve and canine plasma sample dilution comparison

Reagent type: two-step sandwich immunofluorescence detection, streptavidin-coated microplate

Capture antibody: 200ng/well, conjugated biotin

Capture antibody: 200ng/well, labeled europium chelate

Antigen: Canine recombinant NT-proBNP (Cat.#GMP-CAN-NT-proBNP-Ag01)

Sample volume: 50μl

Incubation time: 40 minutes at room temperature

 

Quantitative detection of NT-proBNP in plasma of healthy dogs and dogs with heart disease

These immunoassays have a sensitivity of 22 pg/ml 1 for measuring recombinant NT-proBNP. Figures 1 and 2 display the suggested pairing calibration curves. Conversion of 1 NT-proBNP concentration: 1 pmol/L = 1÷10.545 pg/ml Please be aware that a variety of factors might affect how well an immunoassay test performs. These consist of the labeling procedure, the kind of label coupled to the detection antibody, and the detection platform. Thus, on our customers' immunoassay systems, other NT-proBNP antibody pairs with non-overlapping epitopes could perform better.

 

Figure 3. NT-proBNP concentration in EDTA plasma samples from healthy dogs and dogs with heart disease.

Reagent type: two-step sandwich immunofluorescence detection, streptavidin-coated microplate

Capture antibody: GMP-CAN-NT-proBNP-Ab01, 1μg/well

Capture antibody: GMP-CAN-NT-proBNP-Ab02, 200ng/well, labeled europium chelate

Calibrator: Canine recombinant NT-proBNP (Cat.# GMP-CAN-NT-proBNP-Ag01)

Sample volume: 50μl

Incubation time: 40 minutes at room temperature

 

Improved apparent stability of endogenous NT-proBNP in plasma samples

The gradual breakdown of proteins over time poses a challenge in accurately measuring NT-proBNP content in samples (5-7). Proper sample handling and storage are crucial, but selecting antibodies for detection reagents is equally important. Immunoreactivity decreases if an epitope in either the capture or detection antibodies is broken or located at a protease cleavage point. Therefore, using antibodies that precisely target the stabilizing region can enhance analyte stability. In developing canine NT-proBNP detection reagents, it's vital to choose antibodies resistant to proteolytic degradation of NT-proBNP.

 

Our antibodies were tested for their ability to detect endogenous canine NT-proBNP during sample storage. EDTA plasma from dogs with heart disease was incubated at two temperatures: +4°C and +20°C. At +4°C, NT-proBNP remains stable for at least 72 hours (95–105% of initial immunoreactivity, Figure 4A). Simultaneously, at +20°C, about 98% of the initial immunoreactivity is detectable after 24 hours (Figure 4B). Using our recommended antibody pairings, plasma can be stored at +4°C for at least 72 hours with minimal loss of immunoreactivity. However, at room temperature, there is a slight weakening of the signal during the first 24 hours. Note that individual plasma or serum samples may vary.

Figure 4 , 5. Stability of endogenous canine NT-proBNP in EDTA plasma

(A) Immunoreactivity of NT-proBNP in plasma samples incubated for 24, 48 and 72 hours at +4°C.

(B) Immunoreactivity of NT-proBNP in plasma samples incubated for 4, 8, 24 and 48 hours at +20°C. Collect protease inhibitor-free EDTA plasma; samples are centrifuged, separated, and stored at -70°C before use. EDTA plasma is stored at +4°C or +20°C with the addition of 0.1% NaN3 to prevent bacterial growth. After incubation, samples were stored at -70°C before measurement. The NT-proBNP concentration in plasma was 9 ng/ml (determined by GMP-CAN-NT-proBNP-Ab01 and GMP-CAN-NT-proBNP-Ab02 immunoassays). Detection using GMP-CAN-NT-proBNP-Ab01 and GMP-CAN-NT-proBNP-Ab05 as capture antibodies and GMP-CAN-NT-proBNP-Ab06 and GMP-CAN-NT-proBNP-Ab02 as detection antibodies respectively The system results are depicted in Figure 3.