Thermo Scientific™ Pierce™ Biotin Quantitation Kit, 12 Tests

The Thermo Scientific™ Pierce™ Biotin Quantitation Kit is a well-established colorimetric assay designed to accurately quantify the level of biotinylation in labeled antibodies, proteins, and other bioconjugates. It is based on the use of the HABA-avidin complex, which exhibits a measurable change in absorbance upon displacement by biotin. This simple and efficient method allows researchers to determine the biotin-to-protein ratio in less than 30 minutes using only standard laboratory equipment.

Key Features & Benefits

• Colorimetric Detection
  • Measured at 500 nm using a standard plate reader or spectrophotometer.
  • No need for fluorescence readers or advanced optics.
• Proven Reliability
  • Based on a well-characterized HABA displacement assay, providing reproducible and accurate biotin measurements.
• Convenient Ready-to-Use Reagents
  • Includes pre-mixed HABA-Avidin complex in Thermo Scientific™ No-Weigh™ Microtubes for ease of use.
  • Positive control included: Biotinylated Horseradish Peroxidase (HRP).
• Flexible Format
  • Suitable for both cuvette-based spectrophotometers and 96-well microplate formats.
• Quick Analysis
  • Simple protocol: mix sample with HABA-avidin complex, measure absorbance decrease, and calculate biotin content using extinction coefficients or online calculator.

How the Assay Works

1. Biotin Displaces HABA:
   HABA (4′-hydroxyazobenzene-2-carboxylic acid) forms a weak complex with avidin, resulting in an orange color.
2. Sample Addition:
   When a biotinylated sample is added, biotin competes with HABA for the binding sites on avidin.
3. Color Change & Measurement:
   As HABA is displaced by biotin, the color fades, and the absorbance at 500 nm decreases. This change correlates directly to the concentration of biotin in the sample.
4. Quantification:
   The change in absorbance is used in conjunction with the known extinction coefficient of the HABA-avidin complex to calculate the moles of biotin in the sample.

Kit Contents

• Sufficient for: 12 independent biotinylation measurements

Required Equipment

• Spectrophotometer or plate reader (set at 500 nm)
• Standard cuvettes or 96-well plates

Applications

• Determining biotinylation efficiency in antibodies, enzymes, and other proteins
• Optimizing reagent-to-protein ratios for streptavidin-based assays
• Quality control during protein labeling and conjugation workflows
• Supporting development of biotin-based detection and purification systems

Storage & Stability

• Store entire kit at 4°C
• Kit is shipped at ambient temperature

The Pierce™ Biotin Quantitation Kit provides a fast, reliable, and easy-to-use solution for measuring the biotin content of labeled proteins. It is ideal for researchers needing a cost-effective, colorimetric alternative to fluorescence-based assays, especially in workflows that rely on precise control of biotin conjugation levels. Its simplicity, sensitivity, and flexibility make it a valuable tool for protein labeling QC, assay development, and bioconjugate characterization.