Albumin (ALB) Content Assay Kit (BCG, Micro Method)

Cat. No.: A1505873

GRADE & PURITY BioReagent

Overview

Total Protein (TP) consists of albumin and globulin. Methods for albumin detection include the biuret method, tryptophan method, and dye-binding method. The dye-binding method for albumin can utilize Bromocresol Green (BCG) or Bromocresol Purple (BCP). These dyes have a high affinity for albumin. Typically, the initial rate of dye binding to albumin is monitored, which is proportional to the albumin concentration in the sample.

Assay Principle

Under acidic conditions, albumin molecules carry a positive charge and bind to the negatively charged Bromocresol Green (BCG), forming a blue-green complex. This complex has an absorption peak at 628 nm. The absorbance of this complex is proportional to the albumin concentration. By comparing it with a similarly processed albumin standard, the albumin concentration in the test sample can be determined. This method is commonly used for measuring protein content in human or animal serum, plasma, tissues, etc. It is simple, specific, and can be performed manually or using an automated analyzer. This kit is for research use only and is not suitable for clinical diagnosis or other purposes.

A1505873	Component	100T	Storage
A1505873A	BCG Reagent	25 mL	2-8℃. Store in the dark.
A1505873B	Albumin Standard	20 mg	RT
A1505873C	Albumin Standard Preparation Solution	2 mL	RT
A1505873D	BCG Blank Reagent	10 mL	RT

Required Materials Not Provided

Centrifuge tubes, Small test tubes, Microplate reader, 96-well plate

Experimental Procedure

1. Preparation of Albumin Standard Solution

• Add 0.5 mL of Albumin Standard Preparation Solution (or diluent) to the Albumin Standard (20 mg). Dissolve completely to prepare a 40 mg/mL Albumin Standard Stock Solution.

• The solution can be used immediately after preparation.

• Store the dissolved Albumin Standard Stock Solution at -20°C.

• Important Note: The albumin standard should be dissolved in the same diluent used for the test samples. For example, if the test protein is dissolved in sucrose, dissolve the albumin standard in sucrose. Generally, 0.9% NaCl or PBS can also be used as the diluent for the albumin standard.

2. Sample Preparation

• Serum/Plasma Samples: Use directly, take 5 μL for assay.

• Tissue Samples: Homogenize the tissue in ice-cold physiological saline or PBS at a ratio of 1:9 (tissue weight (g) : saline volume (mL)). Centrifuge the homogenate at 2500 g for 10 minutes. Use 5 μL of the supernatant for assay.

3. Albumin Assay Setup

• Set up Blank, Standard, and Test wells according to the table below.

• Add reagents in the specified order, avoiding bubbles.

• If the sample concentration is too high, reduce the sample volume or dilute appropriately before assaying. Duplicate wells are recommended.

Reagent (μL)	Blank Well	Standard Well	Test Well
Albumin Standard Preparation Solution	5	−	−
Albumin Standard Solution (40 mg/mL)	−	5	−
Test Sample (Serum, Plasma, Tissue Homogenate)	−	−	5
BCG Reagent	250	250	250

4. Albumin Measurement

• Zero the microplate reader with the Blank well.

• Add the BCG Reagent sequentially to all wells and mix immediately.

• Incubate at room temperature for (30 ± 3) seconds.

• Read the absorbance of the Standard well and Test wells at 628 nm (recorded as A_standard and A_test).

5. (Alternative Step) Handling Lipemic/Turbid Samples

• For lipemic or turbid samples, set up a "Sample Blank" well:

  • Add 5 μL of test sample to 250 μL of BCG Blank Reagent. This is the "Sample Blank" well.

  • Zero the instrument with the BCG Blank Reagent.

  • Read the absorbance of the "Sample Blank" well (A_{sample blank}).

  • Use the net absorbance (A_test - A_{sample blank}) to calculate the albumin concentration.

6. Calculation

• Albumin (g/L) = (A_test / A_standard) × Albumin Standard Concentration (g/L)

  • *(Where the Albumin Standard Concentration is 40 g/L if using the prepared stock solution directly in the calculation, but note the standard well contains a diluted amount. Ensure consistency in calculation based on the standard curve or direct ratio as implied. The formula provided assumes a direct proportionality between the absorbance in the test well and the standard well, using the known standard concentration.)*

  • *Clarification: If the standard well contains 5μL of 40 mg/mL standard, this is 0.2 mg of albumin in 255μL total volume. The calculation essentially compares the absorbance produced by the sample to that produced by a known amount of standard under identical conditions.*

Reference Interval

Children (4-14 years)	38～54g/L
Healthy Adults	34～48g/L

Precautions

1. After dissolving the Albumin Standard powder in the Albumin Standard Preparation Solution, the resulting stock solution contains preservatives, which do not affect the assay. This stock solution can be stored long-term at -20°C.

2. If a microplate reader is unavailable, a spectrophotometer can be used. Note that the number of samples that can be assayed per kit may be significantly reduced when using a spectrophotometer.

3. The linear range of this method is 10 - 60 g/L.

4. The inter-assay coefficient of variation (CV) for normal samples using this method is 6.3%.

5. Use reagents promptly after opening to avoid affecting subsequent experimental results.

Storage and Shipping

Storage

Store at 2-8°C,Protected from light,Room temperature

Shipped In

Wet ice

Stability And Storage

Each component has a shelf life of 1 year under corresponding storage conditions. A1505873A: 2-8℃. Store at 2-8℃ long term (12 months). Store in the dark. A1505873B, A1505873C, A1505873D: Store at room temperature long term (12 months).