Human Hemoglobin (Hb) ELISA Kit

The Human Hemoglobin (Hb) ELISA Kit is a quantitative, sandwich-based enzyme-linked immunosorbent assay designed for the detection and measurement of hemoglobin (Hb) in serum, plasma, cell culture supernatants, tissue homogenates, and other human biological fluids. Hemoglobin is a critical oxygen-transporting protein in erythrocytes, and its levels can serve as a research indicator for anemia, hemolysis, inflammation, and organ injury. This assay provides reliable, sensitive results using only 10 µL of sample per test.

Key Features

• Assay Format: Sandwich ELISA
• Detection Range: 0.5 – 15 μg/mL
• Sensitivity: 0.1 μg/mL
• Sample Volume Required: 10 μL
• Detection Wavelength: 450 nm
• Assay Duration: ~2.5 hours
• Storage Conditions: 2–8 °C
• Shelf Life: 6 months
• Precision:
  • Intra-assay CV <10%
  • Inter-assay CV <12%

Assay Principle

This kit utilizes a double-antibody sandwich technique. Microplate wells are pre-coated with an anti-Human Hb monoclonal antibody. Upon sample addition, a second HRP-conjugated anti-Hb antibody binds to the captured antigen. A colorimetric reaction is initiated by TMB substrate, producing a blue color that turns yellow upon addition of stop solution. Absorbance is measured at 450 nm, and Hb concentration is calculated using a standard curve.

Kit Contents

Validated Sample Types

• Serum & Plasma (EDTA/citrate anticoagulated)
• Urine, CSF, Pleural Fluid
• Cell Culture Supernatants & Lysates
• Tissue Homogenates

Note: Do not use samples containing sodium azide (NaN₃), which inhibits HRP. Avoid repeated freeze-thaw cycles.

Assay Procedure Summary

1. Standard Preparation: Serially dilute from 18 → 1 μg/mL.
2. Sample Addition: 40 μL sample diluent + 10 μL sample per well (except blank).
3. Incubation: 30 min at 37 °C.
4. Wash: 5× using diluted wash buffer.
5. Add HRP-Conjugate: 50 μL/well, incubate 30 min at 37 °C.
6. Wash Again, then add 50 μL each of Chromogen A & B.
7. Color Development: Incubate 15 minutes in the dark.
8. Stop Reaction: Add 50 μL stop solution; read OD at 450 nm within 15 minutes.

Performance Characteristics

A log-log standard curve is plotted to determine concentrations. Multiply final values by dilution factor (typically 5) for original concentration.

The Human Hemoglobin (Hb) ELISA Kit (SL2377Hu) provides a reliable, sensitive, and reproducible solution for measuring total hemoglobin in various human biological fluids. Its broad applicability, low detection limit, and minimal sample requirement make it a valuable research tool in hematology, nephrology, toxicology, and clinical pathology. With simple protocols and robust performance, this kit is well-suited for academic, pharmaceutical, and clinical laboratories conducting hemoglobin-related studies.