A method of HbF determination for potential use in underdeveloped countries

Area of Science:

• Biochemistry
• Hematology
• Medical Diagnostics

Background:

• Sickle cell disease management requires monitoring treatment efficacy.
• Hydroxyurea therapy aims to increase fetal hemoglobin (HbF) levels.
• Accurate and accessible HbF determination is vital, especially in underdeveloped countries.

Purpose of the Study:

• To develop a simple, cost-effective method for HbF determination.
• To enable monitoring of sickle cell patient response to hydroxyurea treatment in resource-limited settings.

Main Methods:

• Adapted the Kleihauer-Betke (K-B) test principle using a citrate solution to selectively elute adult hemoglobin (HbA).
• Spectrophotometric absorbance of the eluate was used as the primary outcome measure.
• Optimized procedure parameters including wavelength (395 nm), centrifugation time (30 sec), incubation time (6 min), blood volume (20 microL), and buffer composition (0.07 M sodium citrate in 0.06 M sodium phosphate).

Main Results:

• The developed method demonstrated reproducibility (N=39) with distinct absorbance values for HbA, HbA+F, and HbF.
• Mean absorbance values: HbA = 1.285 (SD 0.069), HbA+F = 0.690 (SD 0.050), HbF = 0.035 (SD 0.005).
• The method exhibited excellent linearity (r² = 0.99).

Conclusions:

• A simple, cost-effective HbF determination method was successfully developed and optimized.
• This method shows significant potential for clinical application in underdeveloped countries.
• It can aid in assessing sickle cell patient response to hydroxyurea therapy.