Download biomedica product listBig Endothelin-1 ELISA | BI-20082H
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Category number
BI-20082H
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Method
Sandwich ELISA, HRP/TMB, 12×8-well detachable strips
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Sample type
Serum, EDTA plasma. citrate plasma, heparın plasma
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Sample volume
50 µl / well
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Assay time
4 h / 1 h / 30 min
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Sensitivity
0.02 pmol/l (= 0.086 pg/ml)
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Standard range
0 – 3 pmol/l (= 0 – 12.85 pg/ml)
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Conversion factor
1 pmol/l = 4.283 pg/ml (MW: 4.283 kDa)
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Precision
In-between-run (n=5): ≤ 5 % CV
Within-run (n=10): ≤ 4 % CV
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Specificity
Endogenous and recombinant Big Endothelin-1 (Big ET-1 (1-38)).
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Cross-reactivity
Human ET1/2/3 (1-21): <1%, human ET2 (1-37): <1%, human ET1/2 (1-38): <1%, human BigET1/2 (22-38) : <1%, human BigET2 (22-37) : <1%,
porcine BigET (1-39): 21%, rat BigET1 (1-39): 10%, Sarafotoxin: <1%
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Validation Data
See validation data tab for: precision, accuracy, diltuion linearity, values for healthy donors, etc.
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Use
Research use only
Big Endothelin-1 ELISA Product Overview
The Big Endothelin-1 immunoassay is a 5.5 hour, 96-well sandwich ELISA for the quantitative determination of Big Endothelin-1 in serum and plasma. The assay employs human serum-based standards to ensure the measurement of biologically reliable data.
Big Endothelin-1 ELISA Assay Principle
The Big Endothelin 1 ELISA kit is a sandwich enzyme immunoassay for the quantitative determination of Big Endothelin-1 (Big ET-1) in human serum and plasma samples.
The figure below explains the principle of the Big et-1 ELISA:
Capture Antibody: polyclonal sheep-anti human Big Endothelin-1 IgG antibody
Detection Antibody: monoclonal mouse anti-human Big Endothelin-1 IgG antibody
Standard: synthetic human Big Endothelin-1 (1-38) in human serum
In a first step, standard/control/sample and detection antibody (monoclonal mouse anti- human Big Endothelin 1 antibody) are pipetted into the wells of the microtiter strips, which are pre-coated with polyclonal sheep anti-human Big Endothelin-1 antibody. Big Endothelin-1 present in the standard/control/sample binds to the pre-coated antibody in the well and forms a sandwich with the detection antibody. After a washing step, which removes all non-specifically bound and unbound material, the conjugate (streptavidin-HRP) is pipetted into the wells and reacts with the detection antibody. After another washing step, the substrate (tetramethylbenzidine, TMB) is pipetted into the wells. The enzyme-catalyzed color change of the substrate is directly proportional to the amount of Big Endothelin-1. This color change is detectable with a standard microtiter plate reader. The concentration of Big Endothelin-1 in the sample is determined directly from the dose response curve.
Big Endothelin-1 ELISA Typical Standard Curve
The figure below shows a typical standard curve for the Big Endothelin ELISA. The immunoassay is calibrated against recombinant Big et1 peptide:
Big Endothelin-1 ELISA Kit Components
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Contents |
Description |
Quantity |
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PLATE |
Polyclonal sheep anti-human Big Endothelin-1 antibody pre-coated microtiter strips in strip holder, packed in aluminum bag with desiccant |
12 x 8 tests |
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WASHBUF |
Wash buffer concentrate 20 x, natural cap |
1 x 50 ml |
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STD |
Standards 1-6 (0; 0.10; 0.20; 0.40; 1; 3 pmol/l), synthetic human Big Endothelin-1 in human serum, white caps, lyophilized |
6 vials |
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CTRL |
Control, yellow cap, lyophilized, exact concentration see label |
1 vial |
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AB |
Monoclonal mouse anti-human Big Endothelin-1 antibody, biotin labeled, red dye, green cap, ready to use |
1 x 18 ml |
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CONJ |
Conjugate (streptavidin-HRP), amber cap, ready to use |
1 x 22 ml |
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SUB |
Substrate (TMB solution), blue cap, ready to use |
1 x 22 ml |
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STOP |
Stop solution, white cap, ready to use |
1 x 7 ml |
Storage instructions: All reagents of the Big Endothelin-1 ELISA kit are stable at 4°C until the expiry date stated on the label of each reagent.
Serum, EDTA plasma, heparın plasma, citrate plasma, cell culture supernatant and urine are suitable for use in this Big Endothelin ELISA kit. Do not change sample type during studies. We recommend duplicate measurements for all samples, standards and controls. The sample collection and storage conditions listed are intended as general guidelines.
Serum & Plasma
Collect venous blood samples in standardized serum separator tubes (SST) or standardized blood collection tubes using EDTA, heparın or citrate as an anticoagulant. For serum samples, allow samples to clot for 30 minutes at room temperature. Perform separation by centrifugation according to the tube manufacturer’s instructions for use. Assay the acquired samples immediately or aliquot and store at -25°C or lower. Lipemic or haemolyzed samples may give erroneous results. Do not freeze-thaw samples more than four times.
Reagent Preparation
Wash Buffer
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1. |
Bring the WASHBUF concentrate to room temperature. Crystals in the buffer concentrate will dissolve at room temperature (18-26°C). |
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2. |
Dilute the WASHBUF concentrate 1:20, e.g. 50 ml WASHBUF + 950 ml distilled or deionized water. Only use diluted WASHBUF when performing the assay. |
The diluted WASHBUF is stable up to one month at 4°C (2-8°C).
Standards & Controls for Serum, Plasma and Urine Measurements
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1. |
Pipette 250 µl of distilled or deionized water into each standard (STD) and control (CTRL) vial. The exact concentration is printed on the label of each vial. |
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2. |
Leave at room temperature (18-26°C) for 15 min. Vortex gently. |
Reconstituted STDs and CTRL are stable at -25°C or lower until expiry date stated on the label. Avoid freeze-thaw cycles!
Sample Preparation
Bring samples to room temperature and mix samples gently to ensure the samples are homogenous. We recommend duplicate measurements for all samples.
Samples for which the OD value exceeds the highest point of the standard range can be diluted with STD 1 or Big Endothelin-1 negative human serum.
Big Endothelin-1 ELISA Assay Protocol
Read the entire protocol before beginning the assay.
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1. |
Bring reagents and samples to room temperature (18-24°C). |
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2. |
Mark positions for STD/CTRL/SAMPLE (standard/control/sample) on the protocol sheet. |
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3. |
Take microtiter strips out of the aluminum bag. Store unused strips with desiccant at 4°C in the aluminum bag. Strips are stable until expiry date stated on the label. |
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4. |
Add 50 µl STD/CTRL/SAMPLE (standard/control/sample) in duplicates into the respective wells. |
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5. |
Add 150 µl AB (biotinylated anti-Big Endothelin-1 antibody, green cap) into each well. Swirl gently. |
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4. |
Cover the plate tightly, swirl gently and incubate for 4 hours at room temperature (18-24°C) in the dark. |
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5. |
Aspirate and wash wells 5 x with 300 µl diluted WASHBUF. After the final wash, remove the remaining WASHBUF by strongly tapping plate against a paper towel. |
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6. |
Add 200 µl CONJ (conjugate, amber cap) into each well. |
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7. |
Cover tightly and incubate for 1 hour at room temperature in the dark. |
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8. |
Aspirate and wash wells 5 x with 300 µl diluted WASHBUF. After the final wash, remove remaining WASHBUF by strongly tapping plate against a paper towel. |
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9. |
Add 200 µl SUB (substrate, blue cap) into each well. |
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10. |
Incubate for 30 min at room temperature in the dark. |
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11. |
Add 50 µl STOP (stop solution, white cap) into each well. |
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12. |
Measure absorbance immediately at 450 nm with reference 630 nm, if available. |
Calculation of Results
Read the optical density (OD) of all wells on a plate reader using 450 nm wavelength (reference wavelength 630 nm). If the OD of the highest STD is outside the measuring range of photometer the plate can be re-measured at 405 nm (correction wavelength 630 nm). Construct a standard curve from the absorbance read-outs of the standards using commercially available software capable of generating a four-parameter logistic (4-PL) fit. Alternatively, plot the standards’ concentration on the x-axis against the mean absorbance for each standard on the y-axis and draw a best fit curve through the points on the graph. Curve fitting algorithms other than 4-PL have not been validated and will need to be evaluated by the user.
Obtain sample concentrations from the standard curve. If required, pmol/l can be converted into pg/ml by applying a conversion factor (1 pg/ml =0.2335 pmol/l (MW: 4.283 kDa)). Respective dilution factors have to be considered when calculating the final concentration of the sample.
The quality control protocol supplied with the kit shows the results of the final release QC for each kit at production date. Data for OD obtained by customers may differ due to various influences and/or due to the normal decrease of signal intensity during shelf life. However, this does not affect validity of results as long as an OD of 1.00 or higher is obtained for the standard with the highest concentration and the control value is in range (target range see label).
Big Endothelin-1 Protein
Big Endothelin-1 (BigET-1) is a peptide of 38 amino acids and is the precursor of Endothelin-1 (ET-1), consisting of amino acids 1-21 (http://www.uniprot.org/uniprot/P05305). ET-1 is a potent vasoconstrictor and is produced by vascular endothelial cells. Accordingly, it has a wide tissue distribution (http://www.ncbi.nlm.nih.gov/UniGene/ESTProfileViewer.cgi?uglist=Hs.511899). The cleavage of BigET-1 by the Endothelin-converting enzyme (ECE) converts BigET-1 to Endothelin (ET) and to a C‑terminal fragment.
The half-life of ET-1 (1-21) in plasma is less than one minute, which may preclude its use as a diagnostic marker, whereas clearance of the precursor peptide BigET-1 is much slower. As Big-Endothelin-1 is produced at amounts equimolar to ET-1 and has higher plasma stability, it is more suitable for diagnostics and monitoring of heart failure (Gergei et al., 2018). Hence, Big ET-1 has the same physiological role as ET-1 while being a more reliable marker.
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Molecular Weight |
4.283 kDa |
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Cellular localisation |
Secreted |
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Post-translational modifications |
Disulphide bonds, post-translational processing |
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Sequence similarities |
Endothelin family |
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Alternative Names |
For endothelin gene: Endothelin 1, Preproendothelin-1, Endothelin-1, PPET1, ARCND3, HDLCQ7, ET1, QME |
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Entrez/NCBI ID |
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Genecards |
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OMIM |
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PDB |
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Pfam |
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Protein Atlas |
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Uniprot ID |
Big Endothelin-1 Function
Preproendothelin, a 203-amino-acid (AA) peptide is encoded by the ET-1 gene. It is processed resulting in Big Endothelin-1 (Big ET-1), containing 38 amino acids. Big-ET-1 is then converted by the endothelin-converting enzyme (ECE) into the biologically active Endothelin-1 peptide (amino acids 1-21). Endothelin-1 circulates in low concentrations and has a very short plasma half-life. Big-Endothelin-1, the precursor is a stable peptide with a plasma half-life of about 30 minutes. Measurement of Big ET-1 reflects Endothelin levels.
ET-1 is secreted, as a paracrine hormone, mainly from endothelial cells towards the smooth muscle cell layer of the vessels, to produce immediate vasoconstriction. Endothelin-1 (ET-1) has in fact been described as the most potent vasoconstrictor substance identified to date, and overactivation or dysfunction of the ET system has been implicated in the development and progression of hypertension in both clinical and experimental studies (Gillis EE et al., 2016). Big endothelin-1 has been demonstrated to be correlated with inflammation, endothelial dysfunction and arterial stiffness (Zhou BY et al., 2017). Both Big Endothelin and Endothelin are strong independent predictors of survival in patients with congestive heart failure and identify a population with very high-risk mortality. Endothelin plays a pathophysiological role in the development of cardiovascular disease (reviewed in Ohkita et al., 2012). ET-1 is also a major factor in the development of chronic kidney disease (CKD) as it contributes to hypertension, proteinuria, and renal inflammation in CKD. Numerous studies have demonstrated that ET-1 directly stimulates inflammation both in the vasculature and in the kidney, and this occurs in the absence of hypertension (Speed and Pollack, 2012).
Endothelins act via activation of two receptor subtypes, ETA and ETB receptors. Endothelin antagonists are currently used clinically in the treatment for patients with pulmonary hypertension (Davenport AP et al., 2016, Houde M et al., 2016), and are considered to have further target diseases as heart failure, cardiac hypertrophy and other cardiac diseases, renal diseases, systemic hypertension, and cerebral vasospasm (Miyauchi T and S Sakai, 2018).
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Cancer
- Oral squamous cell carcinoma (Mankapure et al., 2015)
- Endothelin therapeutics in cancer (Rosanò and Bagnato, 2016)
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Cardiovascular Disease
- Hypertension (Gillis EE et al., 2016)
- Congestive heart failure (reviewed in Ohkita et al., 2012)
- Arterial stiffness (Zhou BY et al., 2017)
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Nephrology
- Renal insufficiency (Fischer et al.,2017)
- During and after graft rejection (Jeremy et al., 2011)
- Chronic kidney disease (Speed and Pollack, 2012)
All Biomedica ELISAs are validated according to FDA/ICH/EMEA guidelines. For more information about our validation guidelines, please refer to our quality page and published validation guidelines and literature.
Validation literature
CPMP/ICH/381/95 ICH Q2(R1) "Validation of Analytical Procedures: Text and Methodology" including:
ICH Q2A "Text on Validation of Analytical Procedures"
ICH Q2B " Validation of Analytical Procedures: Methodology"
Calibration
This immunoassay is calibrated against recombinant human Big Endothelin-1 (1-38).
Big Endothelin-1 ELISA Detection Limit & Sensitivity
To determine the sensitivity of the Big ET-1 ELISA, experiments measuring the lower limit of detection (LOD) and the lower limit of quantification (LLOQ) were conducted.
The LOD, also called the detection limit, is the lowest point at which a signal can be distinguished above the background signal, i.e. the signal that is measured in the absence of Big Endothelin-1, with a confidence level of 99%. It is defined as the mean back calculated concentration of standard 1 (0 pmol/l of Big Endothelin-1) plus three times the standard deviation of the measurements.
The LLOQ, or sensitivity of an assay, is the lowest concentration at which an analyte can be accurately quantified. The criteria for accurate quantification at the LLOQ are an analyte recovery between 75 and 125% and a coefficient of variation (CV) of less than 25%. To determine the LLOQ, standard 2, i.e. the lowest standards containing Big Endothelin-1, is diluted, measured and its concentration back calculated. The lowest dilution, which meets both criteria, is reported as the LLOQ.
The following values were determined for the Big Endothelin-1 ELISA:
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LOD |
0.02 nmol/l |
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LLOQ |
0.03 nmol/l |
Big Endothelin-1 ELISA Precision
The precision of an ELISA is defined as its ability to measure the same concentration consistently within the same experiments carried out by one operator (within-run precision or repeatability) and across several experiments using the same samples but conducted by several operators at different locations using different ELISA lots (in-between-run precision or reproducibility).
Within-Run Precision
Within-run precision was tested by measuring two samples 5 times within one human Big ET-1 ELISA lot. The experiment was conducted by one operator.
|
ID |
n |
Mean Big Endothelin-1 [pmol/l] |
SD [pmol/l] |
CV (%) |
|
Sample 1 |
5 |
0.20 |
0.003 |
2 |
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Sample 2 |
5 |
1.00 |
0.048 |
5 |
In-Between-Run Precision
In-between-run precision was assessed by measuring two samples ten times within three human BigET-1 ELISA kit lots by different operators.
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ID |
n |
Mean Big Endothelin-1 [pmol/l] |
SD [pmol/l] |
CV [%] |
|
Sample 1 |
10 |
0.20 |
0.009 |
4 |
|
Sample 2 |
10 |
1.0 |
0.041 |
4 |
Big Endothelin-1 ELISA Accuracy
The accuracy of an ELISA is defined as the precision with which it can recover samples of known concentrations.
The recovery of the Big Endothelin-1 ELISA was measured by adding recombinant Big Endothelin-1 to human samples containing a known concentration endogenous Big Endothelin-1. The %recovery of the spiked concentration was calculated as the percentage of measured compared over the expected value. All our ELISAs are expected to have %recovery rates within 15% of the nominal value of the sample.
This table shows the summary of the recovery experiments in the Big Endothelin-1 ELISA in different sample matrices:
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% Recovery |
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Sample Matrix |
n |
+ 1 pmol/l |
+ 2 pmol/l |
||
|
Mean |
Range |
Mean |
Range |
||
|
Serum |
14 |
100 |
80 - 127 |
105 |
85 – 119 |
|
EDTA plasma |
3 |
100 |
91 – 108 |
101 |
95 – 114 |
|
Citrate plasma |
3 |
98 |
83 -111 |
105 |
96 - 114 |
|
Heparın plasma |
3 |
97 |
92 – 108 |
102 |
96 – 109 |
Experiments:
Data showing recovery of recombinant Big Endothelin-1 in human serum samples:
|
Big Endothelin-1 [pmol/l] |
% Recovery |
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|
Sample Matrix |
ID |
Reference |
+ 1 pmol/l |
+ 2 pmol/l |
+ 1 pmol/l |
+ 2 pmol/l |
|
|
Serum |
s1 |
0.65 |
1.6 |
2.7 |
97 |
102 |
|
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Serum |
s2 |
0.8 |
2.1 |
2.9 |
117 |
104 |
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Serum |
s3 |
0.02 |
1.3 |
2.4 |
127 |
119 |
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Serum |
s4 |
0.3 |
1.5 |
2.4 |
115 |
104 |
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Serum |
s5 |
0 |
1.1 |
2.5 |
110 |
125 |
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Serum |
s6 |
0 |
0.9 |
2.2 |
90 |
110 |
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Serum |
s7 |
0 |
1.0 |
2.3 |
100 |
115 |
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Serum |
s8 |
0 |
0.8 |
1.7 |
80 |
85 |
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Serum |
s9 |
0 |
0.8 |
1.9 |
80 |
95 |
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Serum |
s10 |
0 |
1.1 |
1.9 |
110 |
95 |
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Serum |
s11 |
0 |
1.0 |
2.1 |
100 |
105 |
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Serum |
s12 |
0.1 |
1.1 |
2.3 |
100 |
110 |
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Serum |
s13 |
0.06 |
0.9 |
1.8 |
85 |
87 |
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Serum |
s14 |
0.06 |
0.9 |
2.3 |
85 |
112 |
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Mean |
100 |
105 |
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Min |
80 |
85 |
|||||
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Max |
127 |
119 |
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Data showing recovery of recombinant Big Endothelin-1 in human EDTA plasma samples:
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Big Endothelin-1 [pmol/l] |
% Recovery |
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Sample Matrix |
ID |
Reference |
+ 1 pmol/l |
+ 2 pmol/l |
+ 1 pmol/l |
+ 2 pmol/l |
|
|
EDTA plasma |
e1 |
0.11 |
1.2 |
2.4 |
108 |
114 |
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EDTA plasma |
e2 |
0.09 |
1.1 |
2.1 |
101 |
100 |
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EDTA plasma |
e3 |
0.1 |
1.0 |
2.0 |
91 |
95 |
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|
Mean |
100 |
101 |
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Data showing recovery of recombinant Big Endothelin-1 in a human citrate plasma sample:
|
Big Endothelin-1 [pmol/l] |
% Recovery |
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Sample Matrix |
ID |
Reference |
+ 1 pmol/l |
+ 2 pmol/l |
+ 1 pmol/l |
+ 2 pmol/l |
|
|
Citrate plasma |
c1 |
0.11 |
1.1 |
2.4 |
99 |
114 |
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Citrate plasma |
c2 |
0.08 |
1.2 |
2.2 |
111 |
106 |
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|
Citrate plasma |
c3 |
0.08 |
0.9 |
2.0 |
83 |
96 |
|
|
Mean |
98 |
105 |
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Data showing recovery of recombinant Big Endothelin-1 in a human heparın plasma sample:
|
Big Endothelin-1 [pmol/l] |
% Recovery |
||||||
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Sample Matrix |
ID |
Reference |
+ 1 pmol/l |
+ 2 pmol/l |
+ 1 pmol/l |
+ 2 pmol/l |
|
|
Heparın plasma |
h1 |
0.11 |
1.2 |
2.3 |
108 |
109 |
|
|
Heparın plasma |
h2 |
0.09 |
1.0 |
2.0 |
92 |
96 |
|
|
Heparın plasma |
h3 |
0.08 |
1.0 |
2.1 |
93 |
101 |
|
|
Mean |
97 |
102 |
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Big Endothelin-1 ELISA Dilution Linearity & Parallelism
Tests of dilution linearity and parallelism ensure that both endogenous and recombinant samples containing Big Endothelin-1 behave in a dose dependent manner and are not affected by matrix effects. Dilution linearity assesses the accuracy of measurements in diluted human samples spiked with known concentrations of recombinant analyte. By contrast, parallelism refers to dilution linearity in human samples and provides evidence that endogenous analyte behaves same way as the recombinant one. For dilution linearity and parallelism are assessed for each sample type and are considered good if the results are within 20% of the expected concentration.
Dilution linearity was assessed by serially diluting human samples spiked with 2 pmol/l recombinant Big Endothelin-1 with standard 1 (human serum containing 0 pmol/l Big Endothelin-1).
The table below show the mean recovery and range of serially diluted recombinant Big Endothelin-1 in serum:
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% Recovery of recombinant Big Endothelin-1 in diluted samples |
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|
Sample Matrix |
n |
1+1 |
1+3 |
||
|
Mean |
Range |
Mean |
Range |
||
|
EDTA plasma |
4 |
110 |
99 – 127 |
104 |
91 - 124 |
Data showing dilution linearity of recombinant Big Endothelin-1 spiked into human blood samples (ref) containing endogenous Big Endothelin-1:
|
Big Endothelin-1 [pmol/l] |
% Recovery |
|||||
|
Sample Matrix |
ID |
Ref |
1+1 |
1+3 |
1+1 |
1+3 |
|
EDTA plasma |
e1 |
2.06 |
1.30 |
0.64 |
127 |
124 |
|
EDTA plasma |
e2 |
2.58 |
1.38 |
0.63 |
107 |
97 |
|
EDTA plasma |
e3 |
2.30 |
1.23 |
0.61 |
107 |
106 |
|
EDTA plasma |
e4 |
2.64 |
1.31 |
0.60 |
99 |
91 |
|
Mean |
110 |
104 |
||||
|
Min |
99 |
91 |
||||
|
Max |
127 |
124 |
||||
Parallelism was assessed by serially diluting human samples containing endogenous Big Endothelin-1 with with standard 1 (human serum containing 0 pmol/l Big Endothelin-1).
The table below show the mean recovery and range of serially diluted endogenous Big Endothelin-1 in human serum:
|
% Recovery of endogenous Big Endothelin-1 in diluted samples |
|||||
|
Sample Matrix |
n |
1+1 |
1+3 |
||
|
Mean |
Range |
Mean |
Range |
||
| Serum | 8 | 90 | 62 - 105 | 96 | 71 - 126 |
Data showing dilution linearity of endogenous Big Endothelin-1 in human serum samples:
|
Big Endothelin-1 [pmol/l] |
% Recovery |
|||||
|
Sample Matrix |
ID |
Ref |
1+1 |
1+3 |
1+1 |
1+3 |
|
Serum |
s1 |
1.36 |
0.42 |
0.24 |
62 |
71 |
|
Serum |
s2 |
0.24 |
0.13 |
0.08 |
105 |
126 |
|
Serum |
s3 |
0.48 |
0.25 |
0.13 |
103 |
110 |
|
Serum |
s4 |
1.46 |
0.64 |
0.33 |
87 |
92 |
|
Serum |
s5 |
0.78 |
0.37 |
0.20 |
94 |
102 |
|
Serum |
s6 |
1.43 |
0.56 |
0.31 |
79 |
87 |
|
Serum |
s7 |
3.11 |
1.47 |
0.59 |
95 |
76 |
|
Serum |
s8 |
2.67 |
1.28 |
0.69 |
96 |
104 |
|
Mean |
90 |
96 |
||||
|
Min |
62 |
71 |
||||
|
Max |
105 |
126 |
||||
Big Endothelin-1 ELISA Specificity
This assay recognizes endogenous (natural) and recombinant human Big Endothelin-1 (Big ET-1).
Cross Reactivity
The factors listed below were assayed for cross-reactivity:
- Human ET1/2/3 (1-21): <1%
- Human ET2 (1-37): <1%
- Human ET1/2 (1-38): <1%
- Porcine BigET (1-39): 21%
- Human BigET1/2 (22-38): <1%
- Human BigET2 (22-37): <1%
- Rat BigET1 (1-39): 10%
- Sarafotoxin: <1%
Sample Stability
The stability of endogenous Big Endothelin-1 was tested by comparing Big Endothelin-1 measurements in samples that had undergone up to four freeze-thaw cycles.
For freeze-thaw experiments, samples were collected according to the supplier’s instruction using blood collection devices and stored at -80°C. Reference samples were freeze-thawed once. The mean recovery of sample concentration after four freeze-thaw cycles is 109%.
|
Big Endothelin-1 [pmol/l] |
% Recovery after 4 freeze/thaw cycles conmpared to 1 freeze/thaw cycle |
||||
|
ID |
1 x |
2 x |
3 x |
4 x |
|
|
s1 |
0.75 |
0.80 |
0.79 |
0.84 |
111 |
|
s2 |
0.15 |
0.14 |
0.12 |
0.17 |
120 |
|
s3 |
0.15 |
0.13 |
0.14 |
0.16 |
103 |
|
s4 |
0.84 |
0.79 |
0.86 |
0.84 |
100 |
|
Mean |
109 |
||||
Samples can undergo at least up to 4 freeze-thaw cycles.
Sample Values
To provide expected values for circulating Big Endothelin-1, a panel of samples from apparently healthy donors as well as samples from patient cohorts with cardiovascular and renal diseases were tested.
A summary of the results is shown below:
|
Big Endothelin-1 [pmol/l] |
|||||
|
Sample Matrix |
n |
Mean |
Median |
Minimum |
Maximum |
|
App. healthy |
41 |
0.22 |
0.09 |
0.01 |
1.58 |
|
Cardio panel |
30 |
0.8 |
0.6 |
0.2 |
2.1 |
|
Dialysis panel |
36 |
1.1 |
0.9 |
0.4 |
3.0 |
It is recommended to establish the normal range for each laboratory.
Matrix Comparison
To assess whether all tested matrices behave the same way in the Big Endothelin-1 ELISA, concentrations of Big Endothelin-1 were measured in serum, EDTA, heparın, and citrate plasma samples prepared from three apparently healthy donors. Each individual donated blood in all tested sample matrices.
A summary table of Big Endothelin-1 levels in various sample matrices is shown below:
|
Big Endothelin-1 [pmol/l] |
|||||
|
Sample ID |
Serum |
EDTA plasma |
Citrate plasma |
Heparın plasma |
% CV |
|
#1 |
0.1 |
0.11 |
0.11 |
0.11 |
7 |
|
#2 |
0.06 |
0.09 |
0.09 |
0.08 |
17 |
|
#3 |
0.06 |
0.1 |
0.08 |
0.08 |
23 |
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