Charles E. Shields

Hemoglobin function in stored blood

Inorganic phosphate which is known to stimulate red cell glycolysis is present in one of the preservatives for storing whole blood, citrate-phosphate-dextrose (CPD), but not the other, acid-citrate-dextrose (ACD). Both of these preservatives for liquid storage were developed before 2,3-diphosphoglycerate (2,3-DPG) was found to be necessary for normal hemoglobin function. In a recent study we have shown that very high concentrations of phosphate (10, 15, and 20 mM) were deleterious for maintaining 2,3-DPG. In the present study a lower range of phosphate concentrations (2, 4, 6, and 8 mM) was studied for maintenance of 2,3-DPG and ATP during storage under blood banking conditions. The lowest concentration, 2 mM, which corresponds to CPD was found to be the best concentration for maintaining 2,3-DPG and thus hemoglobin function. Four mM phosphate was not quite as good but better than no phosphate. Six and 8 mM phosphate were considerably worse.

Clinical Evaluation of Transfused Blood after Long‐term Storage in ACD with Adenine

Whole blood collected in ACD with or without adenine was transfused into patients with various forms of blood loss. The units collected in ACD alone had been stored 14 to 28 days, while those units collected in ACD with adenine had been stored 28 to 42 days. Of the 771 units in the study, 545 were transfused into 295 patients. A total of 276 units were collected in ACD with adenine and were given to 130 patients. Little or no reaction was detected from clinical observation and selected laboratory tests related to the transfusion and/or solution. Isotope labeling of some of the transfused units given to more than 80 patients provided an estimate of the 24‐hour posttransfusion survival and the results were similar to the average survival obtained in normal subjects after single‐unit transfusions. Based on clinical laboratory findings, apparent lack of toxicity combined with adequate support of the bleeding patient suggest that blood stored in adenine is useful after longer storage periods than is blood stored in plain ACD. The 70 per cent 24‐hour survival guideline suggests satisfactory survival after 35 days of storage, and possibly after 42 days.

Erythrocytic abnormalities in experimental malaria.

Summary Studies of erythroeytic indices and osmotic response revealed the presence of 2 abnormal populations of cells in hamsters infected with Plasmodium bergkei. One population consisted of mature nonpar asitized cells which became abnormally small and spherocytic with a diminished cellular surface area. This cell shrinkage could result from the hypersplenism of malaria or possibly from immunologic membrane damage. The other population was composed of parasitized re-ticulocytes which exhibited an increased rate of swelling in hypotonic saline, indicating impairment of the normal restricted cation permeability. The authors wish to thank Mr. Earl H. Fife, Jr., for his critical review of this paper, Mr. Albert E. von Doenhoff, Jr., for assistance with these experiments and Mrs. Sophia Tate for help with preparation of the manuscript.

Biological Alterations Occurring During Red Cell Preservation

Removal of the red blood cell from the normal circulation drastically alters its external environment, depriving the cell of necessary nutrients as well as removing certain protective factors. To counter the loss of nutrient, various blood collecting solutions have been devised in which glucose is used to supply the cell with an energy source. At the same time, attempts are made to reduce cellular metabolism, hence cell demand for energy, usually by using solutions of low pH and/or mechanical factors, that of cooling the blood. The combination of a slower metabolism and a glucose supplement have permitted most storage of red cells in a useful state for perhaps 20–30 days (1,2,3).

Uptake and Retention of Adenine Moiety by Stored Human Red Blood Cells

Tritium‐labeled adenine was added to freshly collected human blood and at selected storage intervals the distribution of the adenine moiety between plasma and erythrocytes was determined. The retention of the radioactive label by erythrocytes was studied by repeated extractions with isotonic saline. The results show that the adenine moiety, which is utilized for net synthesis of various nucleotides, does not appear to be in a steady‐state equilibrium between plasma and red cells. With increasing time of storage, the equilibrium is shifted toward the erythrocytes which reach a maximum radioactive content at 21 days of storage. Specific association of the adenine moiety with red cell populations of different density, investigated by density distribution of erythrocytes with polyester mixtures of known specific gravity, shows that cell populations of low density (young cells) have a high specific activity.

In vivo distribution of injected 14C-dioxyadenine in tissues and organs of normal rats.

A large dose of 2, 8‐dioxyadenine‐14C was injected intravenously into rats and its fate followed at several intervals during the first 24 hours after administration. The distribution of the radioactive material throughout the various tissues or organs was calculated and expressed as a percentage of the injected dose as well as a specific activity per unit organ weight. It appears that dioxy‐adenine is rapidly distributed throughout the organs and tissues of the rats and that specific concentration occurs in a few organs. High retention of the radioactive material and a very high specific activity are observed in the kidney. Smaller but significant concentrations of the 14C‐compound are also observed in muscle and skin, and cannot be attributed to the presence of residual blood. The radioactive material is gradually released from these tissues and, by 24 hours after injection, 92 per cent of the injected dose is found in the urine. Microscopic observation shows deposition of crystals in the kidney. Crystals are also present in the urine and appear as small slightly yellow spheres.

Red cell changes as observed by a simple method for continuously recording osmotic fragility.

Storage of human blood for six to eight weeks in ACD solution resulted in definite hemolysis which paralleled an increase in osmotic fragility determined by a new method. Mechanical injury as a result of delivering blood by airdrop markedly aggravated this hemolysis. Whole blood stored for six weeks in ACD solution with adenine added had less overt hemolysis, less osmotic fragility and better posttransfusion survival than blood from the same subject stored only in ACD solution. Comparison of changes in the osmotic fragility of stored red blood cells showed good correlation with in vivo survival measurements and suggested that this method of testing might be useful in programs to evaluate the effect of additives on the viability of banked blood.

Effect of storage on sheep erythrocytes used in complement studies.

Summary Sheep erythrocytes stored for periods up to 4 weeks in modified Alsevers solution exhibited significant and progressive increases in susceptibility to immune hemolysis in assays conducted with a constant source of human C′. Moreover, the increased susceptibility to immune hemolysis closely paralleled the changes of osmotic fragility and a tendency to undergo spontaneous lysis. The findings suggested an intrinsic relationship between immune hemolysis and osmotic fragility. It was noted that cells stored longer than 3 weeks in modified Alsevers solution could be unduly fragile and hypersensitive to immune lysis thus rendering them unsuitable for serologic tests.

Saliva Agglutinins and Automated Methods for Universal Donor Screening.

Abstract : The objective of the research was to determine the feasibility of automating serum and saliva agglutinins as an additional parameter for group O universal donor screening. The AutoAnalyzer system was employed to automate serum and saliva antibodies. Isoagglutinins A and B, immune A and B antibodies, hemolysins A and B and saliva agglutinins can be screened and quantitated in the AutoAnalyzer. Further experience and evaluation under way is directed toward a better serologic definition of the safe group O universal donor used for A, B, and AB recipients during periods of military exigency and mass casualty situations. (Author)

Evaluation of Automated Multichannel Blood-grouping Apparatus

More than 35,000 blood samples were tested by both automatic and manual methods. The automatic data contained essentially no discrepancies in ABO and less than 0.1 per cent error in Rh blood groups. Mechanical difficulties with the automated system were rare. Obtaining samples was the most common difficulty in mass operation. When cost, number of tests, and time factors are reduced by large numbers of samples, the automatic system provides a useful and accurate method for mass blood grouping. In other respects, the use of automated blood grouping for blood banking will depend upon cost in relation to specific user needs.