Related Experiment Videos
Fluorometric method for phenylalanine microplate assay adapted for phenylketonuria screening
N S Gerasimova1, I V Steklova, T Tuuminen
1All-Union Center for Phenylketonuria, Moscow, U.S.S.R.
Clinical Chemistry
|October 1, 1989
Summary
This study presents a new microplate assay for phenylketonuria screening, offering a reliable, convenient, and cost-effective method for detecting phenylalanine levels in blood spots and plasma.
Area of Science:
- Biochemistry
- Clinical Chemistry
- Neonatal Screening
Background:
- Phenylketonuria (PKU) screening is crucial for early detection and management of the disorder.
- Traditional phenylalanine assays can be labor-intensive and require significant reagent volumes.
Purpose of the Study:
- To adapt the McCaman and Robins fluorometric method for phenylalanine determination into a microplate assay.
- To evaluate the microplate assay's performance for routine PKU screening using plasma and dried blood spots.
Main Methods:
- Fluorometric adaptation of the McCaman and Robins method for microplate format.
- Assay validation using human plasma and dried blood spots.
- Comparison with amino acid analyzer results and the Guthrie inhibition bioassay.
Main Results:
- Achieved sensitivity of 15 mumol/L for plasma and 30 mumol/L for dried blood spots.
- Demonstrated excellent correlation (r = 0.99) with amino acid analyzer results for plasma.
- Exhibited coefficients of variation (CV) less than 10% for both assay types.
Conclusions:
- The microplate assay offers significant advantages in terms of reagent volume, automation, high throughput, and convenience.
- This quantitative method is more reliable, easier to perform, and approximately twofold less costly than the semiquantitative Guthrie inhibition bioassay.
- The developed microplate assay is suitable for routine, high-throughput phenylketonuria screening.