A chemically defined medium containing 18 amino acids, inorganic salts, rhamnose, choline, and ferric pyrophosphate has been developed. The final concentrations of salts and amino acids were modeled after yeast extract. This medium supported the growth of four serogroups of Legionella pneumophila. Growth in shake cultures at 37 degrees C produced a lag time of approximately 5 h and a generation time of 4 h with a maximum growth yield of 10 9 colony-forming units per ml. A soluble brown pigment was observed in the stationary phase of growth. The optimal pH was 6.3. Rhamnose and choline were stimulatory; arginine, serine, threonine, cysteine, valine, and methionine were essential. Supplemental iron was not required to attain maximum growth, but iron deprivation caused an extended lag phase.

A new, automated technique for the preparation of blood components is described. A system of 3 or 4 integrally connected plastic containers (Optipac) is handled by a new type of extractor (Optipress). The container in which the blood is collected has an outlet at the top and another at the bottom. After normal centrifugation to obtain separation of the blood components, these are squeezed out from the top and bottom simultaneously under control of a photocell. The primary separation step results in three components: a leukocyte-poor red-cell suspension in SAGM medium, CPD plasma, and a buffy-coat preparation. The system has been tested in two laboratories (lab A and lab B). A 'heavy-spin' centrifugation to obtain a maximum yield of cell-poor plasma gave the best removal of leukocytes from the red cells; the remaining leukocyte content was 0.46 +/- 0.25 (lab A) and 0.5 +/- 0.4 (lab B) x 10(9)/red-cell unit. Platelet concentrates can be prepared either the normal way via platelet-rich plasma or from buffy coat. Red-cell 24-hour autologous posttransfusion survival using labeling with 51Cr was 87.5 +/- 4.1% (lab A) after 35 days, and 84.2 +/- 4.2% (lab A) and 77.5 +/- 1.5% (lab B) after 42 days. Red-cell morphology and fluidity compared favorably to previous studies using the same additive solution in traditional plastic-bag systems. The total adenine nucleotide concentration was maintained normal for 42 days.(ABSTRACT TRUNCATED AT 250 WORDS)

Red cells stored under blood bank conditions normally show less than 1% spontaneous in vitro hemolysis even after 5 weeks; larger hemolysis may be found if the cells are suspended and stored in a saline-adenine-glucose (SAG) solution with very little trapped plasma. Delay of the addition of the suspension medium, return of 25 ml plasma after a maximal plasma harvest, addition of mannitol 10-30 mmol.1(-1) to the suspension medium were alternative and effective ways of keeping the spontaneous lysis within normal limits. Mechanical traumatization (centrifugation or shaking) caused considerably more damage to the red cells when these were highly concentrated than when they were diluted. A cell suspension in SAG is a more suitable product for hemotherapy than strongly packed red cell concentrates.

The medium described is a simple yeast extract broth capable of growing large number of Legionella neumophila, the causative organism of Legionnaires disease. Filtration was chosen as a means of sterilization, since medium that was autoclaved did not support growth without the presence of Norite A. The filtered medium gave rapid cell growth and maintained the initial antigen production. The observed generation time was 99 min with a maximum cell population of 2 X 10(2) COLONY-FORMING UNITS PER ML IN APPROXIMATELY 40 H.

A new, automated technique for the preparation of blood components is described. A system of 3 or 4 integrally connected plastic containers (Optipac®) is handled by a new type of extractor (Optipress®). The container in which the blood is collected has an outlet at the top and another at the bottom. After normal centrifugation to obtain separation of the blood components, these are squeezed out from the top and bottom simultaneously under control of a photocell. The primary separation step results in three components: a leukocyte-poor red-cell suspension in SAGM medium, CPD plasma, and a buffy-coat preparation. The system has been tested in two laboratories (lab A and lab B). A ‘heavy-spin’ centrifugation to obtain a maximum yield of cell-poor plasma gave the best removal of leukocytes from the red cells; the remaining leukocyte content was 0.46±0.25 (lab A) and 0.5±0.4 (lab B)x 10^9/redcell unit. Platelet concentrates can be prepared either the normal way via platelet-rich plasma or from buffy coat. Red-cell 24-hour autologous posttransfusion survival using labeling with 51Cr was 87.5±4.1% (lab A) after 35 days, and 84.2±4.2% (lab A) and 77.5±1.5% (lab B) after 42 days. Red-cell morphology and fluidity compared favorably to previous studies using the same additive solution in traditional plastic-bag systems. The total adenine nucleotide concentration was maintained normal for 42 days. Storage hemolysis after 6 weeks was slightly higher after a heavy spin, 0.52±0.19% (lab A) and 0.23±0.10% (lab B), than after a light spin, 0.31±0.15 and 0.20±0.09%, respectively. The formation of fibrinopeptide A was very low during the first 2 weeks and then increased to 15±15 nmol/1 after a heavy spin and to 33±11 nmol/1 after a light spin. Kallikrein and spontaneous proteolytic activity increased from day 14 on in the light-spun units but not in the heavy-spun ones. Clinical studies were made in two hospitals. A total of 1,492 transfused blood components prepared with the new system were compared with 1,169 components prepared with a traditional four-container system for making buffy-coat poor red-cell suspensions. No unexpected transfusion reactions occurred. The frequency of febrile reactions was 3/516 (new method) and 2/466 (controls) in a group of patients with predominantly myeloproliferative diseases. The system appears to be a significant improvement for automated preparation of high-quality blood components.

Red blood cells were prepared from CPD whole blood concentrated to 90 per cent hematocrit, diluted with saline-adenine-glucose (SAG) media and then stored for 35 days. The dilution was undertaken to improve the flow properties of the blood and to provide the cells with glucose and adenine allowing prolonged storage. Several ditrerent compositions were tested. ATP could be maintained at about 70 per cent of the initial level and the 24 hour red blood cell posttransfusion survival was 82 per cent. The leakage of potassium was less than in CPDadenine whole blood if the dilution volume was half or less of the red blood cell volume. The only problem was that the hemolysis was greater in SAG-stored blood than in CPD whole blood or undiluted CPD red blood cell concentrate. Hemolysis could be reduced by removal of buffy coat cells or by storage of blood in the presence of synthetic enzyme inhibitors. A chymotrypsin-like enzyme isolated from human leukocytes had a potent hemolytic effect. The hypothesis is presented that red blood cells stored in an electrolyte medium in the presence of leukocytes undergo increased hemolysis because they lack the protecting effect of plasma enzyme inhibitors which normally inhibit any hemolytic enzymes leaking from damaged leukocytes. The study has practical implications for the prolonged storage of bufFy-poor red blood cell preparations to be used in transfusion therapy.

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Blood is normally collected into a combined anticoagulating and preserving medium. We performed a study to ascertain whether improvements could be made by separation of these two functions. Addition of saline-adenine-glucose solutions (40 to 100 ml per blood unit) to buffy-coat-poor red-cell concentrates allowed storage for as long as 35 days with 24-hour erythrocyte post-transfusion survival of 83 +/- 6.8 per cent (+/0 S.D.). Potassium leakage was lower, and in vitro hemolysis somewhat higher than that of whole blood. The microaggregate content after 21 days was 16 per cent of that in whole blood. In over-pressure transfusions the flow rate of red cells was the same with red-cell concentrates to which 80 to 100 ml of suspension medium had been added (hematocrit less than or equal to 60 per cent) as with whole blood. Removal of the buffy coat was essential to reduce hemolysis. We conclude that red cells can be successfully stored in a simple protein-poor medium.