Influence of Chloroquine or Acid Treatment of Human Platelets on the Antigenicity of HLA and the Thrombocyte-Specific' Glycoproteins la/lla, llb, and llb/llla

Neumüller, J.; Tohidast-Akrad, M.; Fischer, M.; Mayr, W.R.

doi:10.1159/000462429

Cited by 3 publications

(7 citation statements)

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“…As has already been shown in a previous study [15], the treatment with citric acid at pH 3 removes the pz-m from the dimeric HLA-class I molecule. Therefore MAB that recognize the whole molecule including the Bz-m bind to platelets treated with citric acid at pH 3 only to a minimal extent, while MAB that recognize only the heavy chain like HC-10 and LA-45 bind strongly to the exposed monomer.…”

Section: Influence Offixation and Thrombin Activation On The Expressisupporting

confidence: 75%

“…It has been argued previously [24] that this 'HLA class I antigen-stripping' is caused by the removal of such HLA class I molecules that are only passively absorbed to the glycocalix of the cell membrane. In contrast, the presence of HLA class I molecules as integral membrane GPs has already been reported by our working group in an earlier publication [15]. In our investigations it was possible to show that immediate fixation ofwhole blood was not appropriate for studies on 'HLA class I antigen-stripping' with citric acid at pH 3.…”

Section: Discussionmentioning

confidence: 38%

“…Protocol A was performed using a diluted flotation medium for mononuclear cells according to our previously described procedure [15]. Protocol B was pcrformed according to the procedure described by Tschope et al [ 161, using differential centrifugation.…”

Section: Pro Hundsmentioning

confidence: 99%

“…The immunolabelling was performed using a 1:10 dilution of MAB. In this dilution, the MAB saturate the binding sites of 10' platelets in 100 pl EDTAiPBS (Ca", Mg" free PBS + 1% BSA and 0.2% EDTA, pH 7.2) as previously shown [15]. Labelling with primary MAB or second-step reagents was carried out for 30 min at + 4 T .…”

Section: Diso-iniinution Ofpiutelets In Whole Bloodmentioning

confidence: 99%

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Standardization of the Flow Cytometric Determination of HLA Class I Antigens, 'Platelet-Specific' Glycoproteins and Activation Markers

Neumüller¹,

Tohidast-Akrad²,

Jilch³

et al. 1995

Vox Sanguinis

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Flow cytometry (FC) provides a reproducible investigation of cell surface antigens on platelets. The aim of this study was to elaborate appropriate protocols and to compare them with other techniques that have already been published. (1) Venipuncture with tubes containing citrate was better for the preservation of the antigenicity than using ACD tubes. The isolated platelets could not be completely distinguished from detritus and protein aggregates. Therefore a platelet concentration between 10^7 and 10^8/ml measurement buffer was necessary to obtain a sufficient resolution by FC. (2) Isolation methods using either differential centrifugation or diluted Ficoll- Hypaque as a flotation medium provided platelets of equal purity. The method with Ficoll- Hypaque resulted in a higher number of isolated platelets than differential centrifugation. The demonstration of platelets and their antigens in whole blood without isolation gave good results provided the platelets were not activated. Activation of platelets with 1 NIH-U thrombin/1 resulted in the loss of a part of the highly activated platelets because of their aggregation. (3) Comparing different concentrations of paraformaldehyde in PBS, fixation with 1% for 15 min provided the best antigen preservation for most of the antigens investigated. Isolation induced platelet activation. In order to avoid this effect, the whole anticoagulated blood was fixed with 1% paraformaldehyde for 15 min immediately after venipuncture. Then the platelets were isolated using diluted Ficoll-Hypaque. In this way, systemic activation of platelets can be detected with antibodies against glycoproteins which are translocated from the a-granules or lysosomes to the cell membrane. These activation markers can be determined on immediately fixed platelets (already in the whole blood) without any interference due to unspecific activation caused by the isolation procedure. (4) Platelet treatment with citric acid at pH 3, in order to remove the antigenicity of HLA-class I molecules, was sensitive to immediate fixation with paraformaldehyde in the whole blood. Fixation after isolating the platelets made it possible to demonstrate antigen stripping, and the free heavy chain, devoid of the β(2)-microglobulin, could be clearly demonstrated. (5) Using standardization beads, the average number of antigenic sites per platelet could be determined for the investigated specificities. It was shown that antibodies which have been directly conjugated or biotinylated and combined with streptavidin-phycoerythrin yielded similar results in terms of the number of antigenic binding sites while unconjugated antibodies in combination with FITC-conjugated anti-mouse-IgG led to overestimation of antigenic binding sites.

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Section: Influence Offixation and Thrombin Activation On The Expressisupporting

confidence: 75%

Section: Discussionmentioning

confidence: 38%

Section: Pro Hundsmentioning

confidence: 99%

Section: Diso-iniinution Ofpiutelets In Whole Bloodmentioning

confidence: 99%

See 2 more Smart Citations

Standardization of the Flow Cytometric Determination of HLA Class I Antigens, 'Platelet-Specific' Glycoproteins and Activation Markers

Neumüller¹,

Tohidast-Akrad²,

Jilch³

et al. 1995

Vox Sanguinis

Self Cite

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“…Aggregation responses were still preserved after acidified CQ treatment [196]. After treatment of platelets with citric acid at pH 3 and CQ, the expression of HLA class I was significantly reduced, while the density of the molecules GPIa/IIa, GPIIb, and GPIIb/IIIa (GP = glycoprotein) carrying thrombocyte-specific antigens was not or only weakly decreased on the surface of the platelets [197]. Pretreatment of the adherent platelets to red cells with CQ diphosphate or citric acid enabled anti-HPA-1a to be used for typing [198].…”

Section: Cq and Removal Of Platelet Membrane Antigensmentioning

confidence: 99%

Antiplatelet and Antileukocyte Effects of Cardiovascular,Immunomodulatory and Chemotherapeutic Drugs

Nosáľ¹

2006

CHAMC

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In vitro and ex vivo interactions of betaadrenoceptor blocking drugs, antihistamines and chloroquine with blood platelets and polymorphonuclear leukocytes resulted in different alterations of regulatory functions of these blood cells. Inhibition of platelet aggregation, arachidonate regulatory pathway, 5-hydroxytryptamine transportation, removal of platelet membrane receptors, inhibition of second messenger pathways at subcellular level and suppression of phagocytosis are indicative of nonreceptor rather than specific receptor interactions. Binding of drugs with biomembranes is reversible depending on the ionic charge of the molecule and hydrophobicity of the bilayer, partition coefficient, pH and pKa of the amphiphilic molecules and other physico-chemical properties of amphiphilic drugs. Alterations in the drug molecule structure alters the drug-phospholipid binding profile. Any change in the metabolism of membrane phospholipids directly or indirectly influences one or more of the important components of the phospholipid-signalling pathway. In addition to changes in phospholipase A, C and D activities, protein kinase C, calmodulin-phosphodiesterase, Ca2+,Mg2+-ATPase, Na+,K+-ATPase and other messengers were found to be changed in cells and tissue after cationic amphiphilic drug (CAD) administration. Although not much has been understood of the mechanism by which some CAD affect immune functions, there are good reasons to suggest that these effects might occur. CADs share sufficient similarities in their structure even though they come from diverse pharmacological classes. CADs affect ion transport, immune functions, tumour growth, serotonin metabolism and several other functions in the body. Extensive therapeutic use and associated side effects have generated a great deal of interest in understanding the nonreceptor interactions with CADs.

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Review: platelet immunology—similarities and differences with red cell immunology

Garratty

1995

Immunohematology

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In many ways platelet and red blood cell (RBC) immunology are similar, but there are important differences. At least 272 RBC antigens have been described. These have been placed into 22 systems (194 antigens), 7 collections (20 antigens), and 2 series (high-frequency antigens [12] and low-frequency antigens [46])1. Platelets express human leukocyte antigens (HLA) (class 1 only), ABH, P (not P1), Lewis, I, and platelet-specific antigens on their membranes. Platelet-specific antigens can be classified into five biallelic systems, originally called PlA1 (Zw), Ko(Sib), Bak, Yuk (Pen), and Br. Each system includes a high-frequency antigen occurring in 96 percent to >99 percent of the population, and homozygosity for a lower-frequency antigen, occurring in 4 percent to <1 percent of the population. In addition, a number of additional low-frequency (private) antigens (e.g., Moa, Caa, Sra, Vaa, Iya, Pea, Groa, Maxa) have been described.2−4 Like RBC antigens and antibodies, platelet-specific antigens and antibodies were first named using letters from the name of the propositus (e.g., Bak, Br, Yuk).

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Influence of Chloroquine or Acid Treatment of Human Platelets on the Antigenicity of HLA and the Thrombocyte-Specific' Glycoproteins la/lla, llb, and llb/llla

Cited by 3 publications

References 7 publications

Standardization of the Flow Cytometric Determination of HLA Class I Antigens, 'Platelet-Specific' Glycoproteins and Activation Markers

Standardization of the Flow Cytometric Determination of HLA Class I Antigens, 'Platelet-Specific' Glycoproteins and Activation Markers

Antiplatelet and Antileukocyte Effects of Cardiovascular,Immunomodulatory and Chemotherapeutic Drugs

Review: platelet immunology—similarities and differences with red cell immunology

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