Leukocyte Agglutination Research Articles

An In Vitro Technic for Quantitating and Studying the Dynamics of Leukocyte Agglutination

Abstract1. An in vitro technic for quantitating leukocyte agglutination has been described.2. With this technic the dynamics of leukocyte agglutination and the response of leukocytes to antigenic agents in both sensitive and nonsensitive systems have been studied.3. The relationship of this study to the problem of agranulocytosis is discussed.

The Demonstration of Two Systems Affecting Blood Leukocytes in the Hypersensitive State with Associated Inflammation

Summary The in vitro agglutination of blood leukocytes has been quantitated. With the quantitative technique, 2 systems affecting blood leukocytes can be demonstrated in the hypersensitive state when associated with inflammation. One system is triggered by the presence of antigen and the other functions in the presence of inflammation. The basic reaction is an agglutination of the leukocytes in the presence of antigen. This reaction is accentuated in the presence of plasma taken from animals in which inflammation was present and is due to an altered reactivity of the leukocytes to the antigen leuko-agglutinating mechanism. Hydrocortisone inhibits in vitro the effect of inflammation but not the effect of antigen.

The in Vitro Quantitative Relationship between Two Systems Affecting Blood Leukocytes in Inflammatory Hypersensitivity Reactions Resulting from Different Antigens

Summary Leukocyte agglutination has been demonstrated to occur in the presence of microgram quantities of antigenic agents in blood from both sensitive and nonsensitive animals. The amount necessary to produce the leuko-agglutinating response is from 10 to 100 times more in blood from nonsensitive animals. The leuko-agglutinating response has been produced with egg albumin, γ-globulin, and pneumococcal polysaccharide. A heat stable, nondializable factor which can produce leukocyte agglutination has been identified in the water soluble serum globulin fraction from animals with inflammation. This factor when diluted 1/2000 to 1/5000 is still able to accentuate and at least in certain instances reduce the threshold of the leuko-agglutinating response to the 3 unrelated antigens. The Inflammatory Leukocyte Agglutinating Factor is inhibited by hydrocortisone, whereas, the action of antigen in the leuko-agglutinating system is not.

Mixed Agglutination of Leukocytes and Erythrocytes in Relation to Studies of Leukocyte Antigens

Abstract Serologic experiments are described in which was studied the agglutination of leukocytes from donors representing various ABO, Rh, and Lewis erythrocyte groups an certain canine blood groups, in the presence of corresponding antisera. Appropriate mixing of antisera with leukocytes and erythrocytes from donors of different groups was seen to produce clumping of leukocytes which did not conform to the reactions of the erythrocytes from the leukocyte donor. When viewed under phase microscopy, certain of these leukocyte clumps, which appeared homogeneous with ordinary light microscopy were found to be clumps of leukocytes mixed with ghosts of erythrocytes reacting with the antibody present. Factors contributing to this apparently immunologically non-specific clumping were the presence of complement-fixing, potentially hemolytic antibody and thermolabile components of serum. A possible relationship with erythrophagocytosis is suggested. These observations indicate that certain results of this and other "leukoagglutination" technics, which have been interpreted as demonstrating the presence of A and B antigens on human leukocytes, deserve re-evaluation and emphasize the importance of developing methods of preparing homogeneous leukocyte suspensions.

Idiopathic Immunoneutropenia: Report of a Case with a Leukocyte Agglutinating Factor in the Serum

Abstract A case report is presented of a woman, aged 49, with persistent neutropenia, increased erythrocyte sedimentation rate, a pathologic serum protein fraction migrating at the same rate as γ-globulin representing about 30 per cent of the total protein, and with undue fatigue as the only symptom. Normal leukocytes, especially neutrophils, were agglutinated by a factor contained in the patient’s serum and bound to the pathologic γ-globulin fraction and active even in a dilution of at least 1:120. The neutropenia may be explained by the formation, in the patient, of antibodies intravascularly agglutinating her own neutrophile leukocytes, which were then eliminated from the circulating blood. The appearance of the bone marrow did not argue against such an assumption. It has not been possible to demonstrate any definite causal factor of the disease. As far as the neutrophile leukocytes are concerned, the disease is analogous with idiopathic immunohemolytic anemia and idiopathic immunothrombocytopenia. No leukocyte agglutination could be demonstrated in sera from two other patients with granulocytopenia or in sera from a number of patients with various diseases and associated increased γ-globulin.

Immunopancytopenia Associated with Incomplete Cold Hemagglutinins in a Case of Primary Atypical Pneumonia

Abstract In a case of primary atypical pneumonia (blood grounp O), incomplete antibodies of the cold agglutinin type with a very wide thermal spectrum (35 to 37 C.) led to a severe immunopancytopenia in the form of severe hemolytic anemia, agranulocytosis, and thrombocytopenia. It is obvious that hemolysis was not induced by cold agglutinins which were active 1:1024 at 4 C. and only 1:32 at 22 C. The indirect cold antibodies which were probably responsible for the hemolysis which occurred were thermolabile (1:16 at 22 C., 1:8 at 37 C.) and could be demonstrated only in the antiglobulin plasma test. During the acute phase, too, a hemolysin of the Dacie type active for normal erythrocytes, but even more active for trypsinized cells, at 37 C. and at a pH of 6.8 and 7.6 could be demonstrated. The severe hemolysis could be stopped by ACTH treatment but the titer of the complete and incomplete cold antibodies showed no distinct change and the direct and indirect Coombs test remained positive for a long time. However, there was a distinct diminution of the thermal amplitude of the incomplete antibodies. A severe agranulocytosis (140 granulocytes) developed which could be improved each time by cortisone. However, permanent improvement resulted only after the disappearance of the incomplete antibodies one year later. A leukopenic factor could be demonstrated by transfusion of 300 cc. blood to a recipient of the same blood group producing a pronounced fall in the granulocytes of from 4200 to 910. Control transfusions from a normal inidividual to the same recipient showed no change and the same is true for the transfusion experiment performed one year later from the now cured patient to the same recipient. Leukocyte agglutination tests were negative, but it is presumed that agglutinins of the incomplete type may have been present. Opsonins could not be demonstrated. The thrombocytopenia never descended below 50,000. The bone marrow showed a distinct shift to the left, i.e. a predominance of unripe forms. On the basis of our previous experimental work (amidopyrine agranulocytosis) it is believed that the changes in the marrow are brought about by a depletion and exhaustion of the bone marrow due to the enormously increased peripheral destruction of the granulocytes by agglutination rather than to an actual inhibition of the marrow. The relation of this immunoleukopenia in our previous findings to the occurrence of leukocyte agglutination in agranulocytosis of allergic origin is discussed. Agranulocytosis and leukopenia may be produced on an immunologic basis in many other cases and thus some form of agranulocytosis may be related to the mechanism of erythrocyte destruction in acquired hemolytic anemia and to the agglutination of thrombocytes ins essential thrombocytopenia on an immunologic basis.

Experimental agranulocytosis. Its production through leukocyte agglutination by antileukocytic serum.

Experimental agranulocytosis. Its production through leukocyte agglutination by antileukocytic serum.

Investigation of leukocyte agglutination in serum of compatible and incompatible blood groups.

Investigation of leukocyte agglutination in serum of compatible and incompatible blood groups.

STUDIES OF BLOOD PASSED THROUGH AN ARTIFICIAL KIDNEY

Abstract 1. In vitro blood studies using the artificial kidney are discussed in detail. 2. In vitro experiments in which samples of blood in cellophane tubing are rotated on the drum showed a diminution in the red cell volume, leukocyte and platelet counts. Dilution, caused by the passage of the dialyzing fluid through the cellophane membrane and by the presence of some of the rinsing fluid, and rotation of the drum are factors of importance in this regard. 3. The influence of heparin and the chemotactic role of cellophane on the leukocytes are discussed. Heparin in concentration of 1 mg./10 cc. of blood causes in vitro agglutination and disintegration of leukocytes after approximately one hour. Photographs are shown that demonstrate coating of the cellophane by granulocytes. This is in keeping with the findings of Chambers and Grand that cellophane exerts a positive chemotactic influence on the granulocytes. 4. Hypersegmentation of the leukocytes was noted in samples of blood rotated on the drum for fifteen minutes. Exteriorization and slow speed centrifugation probably account for this phenomenon.

HYPERHEPARINEMIA, WHITE EMBOLI AND LEUKERGY

ABSTRACT To the Editor:— With reference to Alfred L. Copley's article, Hemorrhagic Diathesis in Hiroshima, The Journal (137: 145 [May 8] 1948), I should like to make the following suggestions:In a number of experiments conducted in our laboratories at the beginning of 1948 I had definite evidence which convinced me that heparin in vitro and in vivo (even in doses smaller than needed for the prevention of blood coagulation) causes agglutination of thrombocytes—thus contradicting the opinion hold by most writers (Jorpes, Solandt and Best, Boyd, Whitby and Britton and others). I am glad that the findings of Copley and his co-workers are the same. Independently of the agglutination of thrombocytes, however, heparin causes also a clumping of the white blood cells, mainly granulocytes. It can be proved that this agglutination of leukocytes is independent of the presence of platelets, because it takes place also in the blood of an

Factors Influencing Leukoagglutination by Antileukocytic Sera

Summary A rapid procedure for leukoagglutination by specific antisera and conditions which influence it are presented. The procedure consists in suspending leukocytes in a small volume of saline, counting the cells with pipette and chamber and measuring out 27,000 cells per mm3 in 0.05 ml of normal saline. To the cells is added 0.5 ml of antiserum diluted with normal saline. The cell-antiserum mixture is centrifuged for one minute and the results are read. The entire procedure is carried out at 22 to 26 C. Agglutination of leukocytes is inhibited at 8 C and the velocity is reduced at 37 C. The optimal temperature is 22 to 26 C. Centrifuging results in the greatest speed of antigen-antibody reaction. Immature leukocytes and especially those outside the body for 2 to 3 days are destroyed more readily than mature cells by hypotonic salt solution. Agglutination requires the presence of intact cells. Electrolytes are necessary for the visible phase of agglutination and the optimal range is 0.85 to 2.5 per cent. Mature leukocytes are agglutinated in lower concentrations. Ageing of cells outside the body interferes with agglutination and mature leukocytes are agglutinated up to the fourteenth day. Immature cells show little or no clumping after the fourth day. Repeated washing of leukocytes does not interfere with agglutination until after the tenth time when most of the cells are destroyed. Hemoglobin up to 50 per cent does not interfere with agglutination. A definite number of leukocytes is necessary for a satisfactory agglutination. Excess or decrease in the number of cells modifies the degree of clumping.