It's about time: The development and validation of a rapid optimized single antigen bead (ROB) assay protocol for...

Area of Science:

• Immunology
• Transplantation Science
• Clinical Diagnostics

Background:

• The LABScreen single antigen bead (SAB) assay is crucial for monitoring human leukocyte antigen (HLA) antibodies in transplant patients.
• Assay turnaround time is critical, particularly for deceased donor selection and post-transplant monitoring.

Purpose of the Study:

• To develop and validate a Rapid Optimized SAB (ROB) protocol.
• To significantly reduce the assay time of the One Lambda LABScreen SAB (OLSAB) procedure without compromising accuracy or sensitivity.

Main Methods:

• Modified the OLSAB protocol by shortening centrifugation cycles and reducing serum and secondary antibody incubation times.
• Increased secondary antibody concentration to compensate for reduced incubation.
• Validated the ROB protocol against the OLSAB protocol using 58 sera samples across two HLA laboratories.

Main Results:

• The ROB protocol reduced assay time from 85 minutes to 25 minutes, a >70% decrease.
• Excellent correlation (r² > 0.98) was observed between ROB and OLSAB protocols for both class I and class II HLA antibodies.
• The ROB protocol showed a trend towards improved inter-laboratory median fluorescence intensity (MFI) concordance compared to OLSAB.

Conclusions:

• The Rapid Optimized SAB (ROB) protocol offers a significantly faster alternative to the standard OLSAB procedure.
• The ROB protocol maintains assay quality and sensitivity, making it suitable for clinical use.
• Implementation of ROB can expedite HLA antibody testing and potentially enhance assay reproducibility.