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A European workshop to investigate technical problems of using HLA monoclonal antibodies in microcytotoxicity assays
Abstract:
Various methods for controlling carryover from monoclonal antibodies (MCA) used in the microcytotoxicity assay to detect HLA antigens were investigated by 45 European laboratories. Carryover was demonstrated in most laboratories when using standard techniques (i.e. as for allo-antisera). Taking into consideration factors such as technical ease, time, accuracy and reagent availability, nearly half the participants selected the "no-touch" technique for adding cells and complement as being the best method for controlling carryover in their laboratory. (This method allows the needle of the Hamilton syringe to touch the oil but not the antibody in each well). Most participants (75%) thought that at least one diluent well was still necessary between each MCA well, even when using their preferred technique.
Insights
To prevent carryover in HLA antigen detection using monoclonal antibodies (MCA), the "no-touch" technique for adding cells and complement is preferred by many labs. This method, along with diluent wells, helps ensure accurate microcytotoxicity assay results.
Area of Science:
- Immunology
- Histocompatibility
Background:
- Monoclonal antibodies (MCA) are crucial for detecting human leukocyte antigen (HLA) antigens via microcytotoxicity assays.
- Carryover between wells in these assays can compromise HLA typing accuracy.
- Standard techniques often fail to adequately control for MCA carryover.
Purpose of the Study:
- To investigate and compare various methods for controlling MCA carryover in HLA microcytotoxicity assays.
- To identify the most effective and practical carryover control technique among European laboratories.
Main Methods:
- A survey of 45 European laboratories was conducted to assess different carryover control strategies.
- Participants evaluated methods based on technical ease, time, accuracy, and reagent availability.
- The "no-touch" technique, where the Hamilton syringe needle touches the oil but not the antibody, was specifically examined.
Main Results:
- Carryover was prevalent in most laboratories using standard microcytotoxicity assay techniques.
- Nearly half of the participating laboratories identified the "no-touch" technique as the optimal method for controlling MCA carryover.
- A significant majority (75%) of participants still considered the use of at least one diluent well essential between MCA wells.
Conclusions:
- The "no-touch" technique offers a practical and effective solution for minimizing MCA carryover in HLA typing.
- Implementing diluent wells remains a recommended practice, even when employing advanced techniques like the "no-touch" method.
- Standardization of carryover control methods is vital for reliable HLA antigen detection.