(h Albumin ELISA, Strip Plate)
The Albuwell Hu kit contains all of the components and reagents you need to assay human albumin in up to 80 duplicate urine or cell free liquid samples (plasma, serum, cell culture supernatant, etc…) for the quantitative
measurement of microalbuminuria in human specimens.
Intended Use: Albuwell Hu is a direct enzyme linked immunosorbent assay (ELISA) for the quantitative determination of albumin in human urine. It is intended for research applications; it is not intended for diagnostic use.
Kit Contents: Your Albuwell Hu kit should contain the following items:
1. 2 Albuwell Assay Plates
2. 2 NHEBSA (Diluent)
3. Human Serum Albumin (hSA) Standard
4. 2 Anti-human Albumin Ab-HRP Conjugate
5. 2 Color Developer
6. 2 Color Stopper
hSA Standard, NHEBSA, Rabbit anti-human Albumin Ab-HRP Conjugate preparations contain 0.05% Proclin 300 (active components
isothiazolones) as preservative. Color Stopper contains dilute (2.0 N) sulfuric acid
Albuwell plates are precoated and ready to use. All kit reagents are supplied in ready to use liquid form. A provision to wash the plates should be made. Tap water has been shown to be suitable in experimental and quality control contexts, but an EIA Wash Buffer with composition: 0.15 M NaCl, 0.01 M triethanolamine (pH 6.8), 0.05% Tween 20 and 0.05 % Proclin 300 (preservative may be
omitted if the buffer is freshly prepared) may be used where tap water is unavailable or proves unsuitable.
Micropipettors capable of delivering 10, 50, 100 and 120 uL are required. Multi-channel pipettors capable of delivering 50 and 100 uL are recommended. In addition, small test tubes are required to complete dilutions (microfuge tubes work well in this application). Finally, a microplate reader equipped to determine absorbance at 450 nm is required.
Technical Background: Albuwell Hu is intended for use as an in-vitro tool for
assessing kidney function in humans. It is simple to perform and highly specific for human albumin. It is a competitive antibody capture ELISA completed in a direct mode.
To that end, the anti-albumin antibody (Ab) is conjugated to horseradish peroxidase (HRP), i.e. directly labeled. To complete the assay, sample and antihuman albumin Ab-HRP are added to human albumin-coated well. The antibody interacts and binds with the albumin immobilized to the stationary phase or with that in the fluid phase, hence the notion of competitive binding.
Washing removes Ab-HRP-Albumin and unbound reactants of the fluid phase from the well. Only the antibody-HRP conjugate that was bound to the albumin of the stationary phase remains, and this is detected using Tetramethylbenzidine (TMB) in a chromogenic reaction. The reaction is stopped with acid, and absorbance is measured at 450 nm.
Absorbance is inversely proportional to the logarithm of albumin in the fluid phase. Specimen Collection and Storage: Collect samples without preservative, and clarify them by centrifugation if necessary. Store clarified urine at 4oC for up to 1 week or at -60oC for up to 2 months. Prior to assay, allow the samples to come to room temperature. Do not apply heat to thaw frozen samples.
Assay Procedure: Allow reagents and samples to come to room temperature before running the assay.
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