Acidified Chloroquine Treatment for the Removal of Class I HLA Antigens

Abstract

With the aim of reducing the damage to platelets while effectively removing class I HLA antigens from their surfaces, we developed a new method using acidified chloroquine diphosphate. Platelets were treated with a 0.2 M solution of chloroquine diphosphate (pH 4.0). More than 90% of the platelets remained viable after treatment. While a marked reduction in reactions of acidified chloroquine-treated platelets with multispecific HLA antisera was noted in comparison with phosphate-buffered-saline-(PBS)-treated platelets, reactions with platelet-specific antibodies were preserved. This was demonstrated by immunofluorescence tests and solid-phase and monoclonal antibody immobilization of platelet antigen assays. Aggregation responses, though reduced in comparison with PBS-treated platelets, were still preserved after acidified chloroquine treatment. Ultrastructural analysis did not show any significant difference from PBS-treated platelets. We conclude that treatment of platelets with acidified chloroquine diphosphate is a simple and effective method for removing class I HLA antigens from their surfaces with minimal damage to their structure and function.

Slot blotting and flow cytometry: two efficient assays for platelet antibody screening among patients with platelet refractoriness

“…Achieving an adequate count increment after transfusion of platelets is a challenging issue that may significantly improve the patients' responsiveness and outcome. Optimal post-transfusion response depends on antibody screening for patients facing frequent platelet intake [2,25]. Currently, a variety of methods, with different pros and cons, have been suggested.…”

“…Regarding the HLA/HPA discrimination, previous studies offer a promising option for HLA stripping and recommend that the treatment of platelets with chloroquine solution can yield HLA-free cells [18,24]. On the other hand, Srivastava and coworkers proposed that acidified chloroquine treatment through a configurational change of membrane glycoproteins and also HLA class I antigen removal can slightly improve the number and accessibility of available sites for HPA [25]. Accordingly, in the present study chloroquine-treated platelets were incubated with selected sera from the patients for whom MAIPA assay was positive, and then, the results of the SB and FC techniques were compared.…”

Antenatal Screening for HPA‐la by Flow Cytometry

“…The platelet phenotyping conducted by the Platelet Immunology Laboratory at the NSW Blood Transfusion Service used a modified solid phase red cell adherence (SPRCA) method ( 12). Genotyping was performed by the polymerase chain reactionallele specific restriction enzyme (PCR-ASRA) method to clarify equivocal results ( I 3).…”

Progress and challenges in platelet serology

“…Unfortunately, they do not allow discrimination between HPA and HLA antibodies. Treatment of platelets with acidified chloroquine diphosphate seems to be a simple method to remove HLA class I antigenicity from test platelets . In case of strong reactive HLA antibodies, however, positive reaction often still remains.…”