Butanol extraction of serum and urinary gamma-glutamyltransferase and its application in clinical diagnosis.

Gamma-glutamyltransferase (gamma-GT) activity was measured in serum and urine before and after extraction with n-butanol. Residual gamma-GT activity after extraction of normal serum with butanol was more than 75%, whereas for normal urine the figure was less than 68%. The loss of activity from urine was not affected by prior dialysis, and the stabilisation which resulted from adding bovine serum albumin at a concentration of 40 g/l did not approach the residual activity with normal serum. In patients with a variety of renal diseases, residual gamma-GT activity in the urine after butanol extraction was inversely correlated with the creatinine clearance. Butanol extraction was performed on serum samples from 182 patients with a variety of diseases. Eighty-one per cent of patients with elevated gamma-GT activities caused by hepatobiliary disease had an increased loss of activity after butanol extraction. By contrast, only 34% of patients with increases gamma-GT activities in whom there was no clinical or other biochemical evidence of hepatic disease, had increased loss of gamma-GT activity after butanol. The reasons for differences between urinary and serum gamma-GT in response to butanol, and the implications in interpreting serum gamma-GT activities are discussed.     

SERUM AND URINARY LYSOZYME (MURAMIDASE) IN MONOCYTIC AND MONOMYELOCYTIC LEUKEMIA

Markedly increased quantities of lysozyme have been found in the serum and urine (ranging to 2.6 g per day) of ten consecutive cases of monocytic and monomyelocytic leukemia. The enzyme has been isolated from the urine of several cases and physicochemically and immunochemically characterized. It is apparently identical to the lysozyme of normal tears, saliva, leukocytes, and serum, but structurally different from the lysozyme of hen's egg white. The activity of the human enzyme assayed with M. lysodeikticus organisms is 3 to 12 times greater than egg white lysozyme at equivalent concentrations. An agar plate method has been developed for quantitating lysozyme activity in small samples (approximately 25 µl) of serum, urine, or other biological fluids. The range and reproducibility of this method were found to be superior to previously available lysozyme assay procedures. Present evidence indicates that lysozyme is the principal, if not the sole, product of the proliferating monocytes in monocytic and monomyelocytic leukemia, and quantitation of serum and urine lysozyme should be a useful diagnostic procedure for these leukemias.     

Biochemical tests in the diagnosis of nephrotoxicity: assessment of the significance

In order to identify early signs of nephrotoxicity in 38 patients with malignant tumors of the testicle, the data of 3 biochemical tests (measurements of gamma-glutamyltranspeptidase (gamma-GT) activity in the urine, of the content of creatinine and urea in blood serum) were compared. Activation of gamma-GT in the urine may be viewed as the first and early sign of nephrotoxicity, since the rise of creatinine and urea in the blood was observed later. The magnitude of gamma-GT activity (means + 2 sigma) - 5.9 Units/mmo le Cr may provide basis for the judgement about nephrotoxicity according to the scheme applied. The fall of gamma-GT activity without any further increase after multiple courses of chemotherapy is of prognostic significance for its evidences profound injury to the renal parenchyma and may serve as basis for correcting the doses of chemotherapeutic drugs. The reduction of gamma-GT activity to less than 3 Units/mmole creatinine (means - 2 sigma) may be regarded as indication for instituting disintoxication therapy.     

Inactivation of endotoxin by a humoral component. II. Interaction of endotoxin with serum and plasma

A humoral substance which inactivates endotoxin in vitro has been shown to be clearly distinguishable from complement, properdin, and specific antibody. For the present, it is designated "endotoxin-detoxifying component" or EDC. Animal species could be grouped in three categories with regard to the EDC activity of their sera; rat serum was highly potent; chimpanzee, dog, horse, and guinea pig sera were much less active; mouse, rabbit, and sheep sera exhibited no activity. The EDC potency of human sera varied widely, ranging from high to barely discernible activity. In contrast to the variations of EDC potency in serum, citrated plasma from all species manifested high potency of about the same magnitude. The influence of time, temperature, pH, and concentration of reactants on the inactivation of endotoxin by EDC was examined. EDC activity in plasma and serum was found to be labile to beating at 56°C. for 1 hour. Bacterial endotoxins, derived by different isolation procedures from smooth and rough Gram-negative species, varied considerably in susceptibility to EDC action.     

THE SURVIVAL OF LEPTOSPIRA (SPIROCH?TA) ICTEROH?MORRHAGI? IN NATURE; OBSERVATIONS CONCERNING MICROCHEMICAL REACTIONS AND INTERMEDIARY HOSTS

1. Leptospira icterohæmorrhagiæ is unable to grow in the urine, either with or without the addition of suitable culture ingredients, the acidity of the urine being detrimental to the growth. It survives less than 24 hours, unless the urine is neutralized or slightly alkalized, when the period of survival is somewhat longer. If suitable nutrient ingredients are added to the neutralized or slightly alkalized urine, the organism is able to grow for about 10 days, after which multiplication ceases. 2. Feces from normal or jaundiced persons destroy Leptospira icterohæmorrhagiæ within 24 hours when a rich culture is added and the mixture allowed to stand at 26°C. The addition of blood serum and corpuscles does not prevent the destruction of the organism. Autoclaved specimens and filtrates of unheated feces do not constitute a suitable medium in which to keep the organism alive for any length of time, but the addition of blood corpuscles and serum in adequate quantities renders them fairly satisfactory as media. Under natural conditions Leptospira icterohæmorrhagiæ cast off in the feces cannot survive more than 24 hours. 3. Polluted water, sewage, and soil will not serve to keep Leptospira icterohæmorrhagiæ alive for more than 3 days at the most. When deprived by filtration or autoclaving of their bacteria they become indifferent diluents and may be used to make up a culture medium when mixed with serum and citrate plasma of a suitable animal. Sterilized soil with a neutral reaction, when added to a culture, has an unfavorable effect upon the growth of the organism. 4. Most of the aerobic bacteria found in feces, sewage, soil, and tap water inhibit the growth of Leptospira icterohæmorrhagiæ when inoculated into the same medium. Bacillus fæcalis alkaligenes and many strains of non-hemolytic streptococci caused the least interference, although growth was never so vigorous or lasting in the media in which they were present as in the control media. Certain pathogenic bacteria (Bacillus typhosus, Bacillus paratyphosus, Bacillus dysenteriæ, pneumococcus) are antagonistic to the growth of the spirochete. 5. Leptospira icterohæmorrhagiæ is highly sensitive to the destructive action of bile, bile salts, and sodium oleate, but resists the action of saponin. In this last respect it differs from many so called spirochetes. The destructive action of these agents is counteracted by blood serum. 6. The larvae and adults of the Culex mosquito, the larvæ of the house-fly and bluebottle fly, wood ticks (Dermacentor andersoni), and leeches failed to become carriers of the spirochetes when fed on infected guinea pigs or their organs; that is, they cannot play the part of an intermediary host of Leptospira icterohæmorrhagiæ.     

The gamma-glutamyltransferase (GGT) isoenzymes in human serum (author's transl)]

With this report we communicate the results of a method for the fractionation of the isoenzymes of gamma-GT in human serum based on a modified technique of Hetland et al., using cellulosa acetate. With this method we observed the appearance of a complex of four band representing: gamma-GT1 = prealbumin-albumin, gamma-GT3 = alpha 1-globulin, gamma-GT3 = alpha 2-globulin, gamma-GT4 = beta-globulin. It has been noted that the gamma-GT2 is the only fraction which constantly appears. This isoenzyme also corresponds to the form, fisiologically present in serum and is presumably preduced in the cells of the bileduct. In the results we observed the appearance of gamma-GT1 only in the course of processes involving certainly cholestasis. This fraction was once reported by other AA., but it has an unknown origin. It was not possible to correlate the appearance of the fractions gamma-GT3 and gamma-GT4 with any specific pathology alterations even though Hetland et al. have affirmed that the gamma-GT3 band derives from hepatic parenchyma.     

THE EFFECT ON SUBSEQUENT AGGLUTINATION OF THE EXPOSURE OF BACTERIA TO HEATED ANTISERUM

It is shown that when dilute rabbit serum rich in agglutinin for the hog cholera bacillus is heated at 75°C. for 20 minutes and the bacilli incubated with the heated serum, agglutination fails to result on the addition of unheated immune serum. When the immune serum is first heated to 80°C., it no longer greatly inhibits secondary agglutination when the organisms are exposed to fresh agglutinin. The abortion bacillus agglutinin acts in a similar manner except that the immune serum must be heated above 80°C. for 20 minutes to prevent the second agglutination. The reactions are specific since control experiments with normal rabbit serum heated at various temperatures failed to influence further agglutination. It has also been shown by precipitation tests that there is definite fixation of serum proteins and bacterial cells with the heated sera which would prevent subsequent agglutination. Furthermore, heated antiserum which would prevent the secondary agglutination still possessed the property of deviating complement in a hemolytic series.     

THE PREPARATION AND PROPERTIES OF INFLUENZA VIRUS VACCINES CONCENTRATED AND PURIFIED BY DIFFERENTIAL CENTRIFUGATION

Influenza virus vaccines containing from 1 to 10 mg. of virus materials per cc. concentrated and purified from infectious allantoic fluids by means of one or two cycles of differential centrifugation and inactivated by different treatments have been prepared and subjected to laboratory tests. Suitable inactivation of the virus preparations with retention of full red cell agglutinating activity and immunizing potency in mice was achieved by treatment with minimal amounts of formaldehyde or ultraviolet light. Treatment with phenol or chloroform failed to cause adequate loss of virus activity. Excessive amounts of formaldehyde or of ultraviolet light were found to cause a loss in red cell agglutinating activity and in immunizing potency. Freezing resulted in the immediate loss of red cell agglutinating activity of the formalinized vaccine. Storage of the vaccines in the frozen state was accompanied by a gradual decrease in red cell agglutinating activity. Drying of the vaccines from the frozen state resulted in a loss of red cell agglutinating activity and, in the case of the formalinized vaccine, in a loss in immunizing potency. There appeared to be at least a rough correlation between red cell agglutinating activity and immunizing potency. The immunizing potency and red cell agglutinating activity of a purified formalinized vaccine containing 2 mg. of virus material per cc. were unchanged following 2 months'' storage at 4° but were measurably decreased following storage for 2 months at 18 to 25° and at 37°. At equivalent dosages of virus material the immunizing potency of formalinized centrifugally purified virus, of formalinized virus purified by the red cell elution method, and of infectious allantoic fluid was not measurably different. The immunizing potency of a formalinized polyvalent vaccine containing centrifugally purified Lee, PR8, and Weiss influenza virus materials at concentrations of 5, 2.5, and 2.5 mg. per cc., respectively, was found to be essentially the same as that of a similar vaccine prepared commercially. In both cases the protection afforded against the Weiss strain appeared to be better than that against the Lee and PR8 strains. The commercially prepared vaccine is being subjected to clinical tests in man at dosage levels ranging from 0.01 mg. to 10 mg. The latter corresponds to a level approximately 100 times that of infectious allantoic fluid. It was found that the bacterial contamination that frequently accompanies operation on a large scale can be controlled by the addition of one part per 10,000 of formalin plus one part per 100,000 of phenyl mercuric nitrate to the allantoic fluid immediately following harvesting, without affecting the quality of the vaccine. This procedure and the use of virus materials purified and concentrated by a single cycle of differential centrifugation by means of the Sharples centrifuge were found to be suitable for the production of influenza virus vaccines on a large scale. By means of this method influenza vaccines possessing 20 or more times the immunizing potency of infectious allantoic fluid and 10 or more times the immunizing potency of the usual commercial vaccine prepared by the red cell elution method can be manufactured rapidly on a very large scale with considerable ease and efficiency.     

STUDIES ON THE BIOCHEMICAL,BIOPHYSICAL, AND IMMUNOGENIC PROPERTIES OF JAPANESE B TYPE ENCEPHALITIS VIRUS AND VACCINES

Studies on the biochemical, biophysical, and immunogenic properties of Japanese B type encephalitis virus and vaccines have been made in order to determine whether a purified vaccine suitable for human use could be obtained by means of differential centrifugation of extracts of infected mouse brains. Studies were also made on extracts of normal mouse brains, it was found that extracts of normal as well as of infected mouse brains contained fairly large amounts of several components of high molecular weight. Components having sedimentation constants near 5 and 40 Svedberg units were found in extracts of infected brains. However the rates of sedimentation of the different components were so similar that it was found impossible, from a practical standpoint, to secure a vaccine consisting largely of virus by means of differential centrifugation. It was also found that a considerable portion of virus was lost or destroyed in the centrifugation process so that it was impossible to secure an effective degree of concentration of immunogenic potency. Although vaccines possessing about twice the immunogenic potency of the starting material were obtained, it was concluded that it was not practical to purify and concentrate Japanese B type encephalitis virus in infected mouse brain extracts by means of differential centrifugation for the production of a vaccine on a large scale. The optimum pH stability range of Japanese B type encephalitis virus activity was found to be near pH 8.5. The virus is inactivated fairly rapidly at pH 7 and very rapidly at more acid reactions. The virus is inactivated rapidly near pH 10. Extracts of infected mouse brains with buffers near pH 8 containing disodium phosphate were found to possess slightly higher titers than saline extracts near pH 7. However vaccines prepared from such extracts were found to possess essentially the same immunogenic potency, hence, although extraction at the more alkaline reaction may perhaps remove more active virus, there was no indication that more immunogenic material was removed at the more alkaline reaction. The use of different diluents in the titration of virus activity and the use of different agents in the preparation and storage of virus suspensions were investigated. It was found that low titers were obtained when Ringer''s solution, phosphate buffer at pH 7, or saline-phosphate buffer at pH 8.2 were used as diluents but that high titers were obtained when 10 per cent rabbit serum in saline or in phosphate buffer, 10 per cent skim milk in saline or in phosphate buffer, or 1 per cent arginine at pH 8.3 were used. Undiluted skim milk adjusted to pH 8.4 was found to be as satisfactory as undiluted rabbit serum for the preparation of infected brain suspensions for storage at –70° C. A satisfactory neutralization test was conducted with virus stored in undiluted skim milk at –70° C. and subsequently diluted with 10 per cent skim milk in saline. The demonstration that skim milk can be substituted for rabbit serum in the storage, titration, and neutralization tests of Japanese B type encephalitis virus is of practical importance, for skim milk is more convenient to prepare and more readily available in many localities.     

A POTENT ANTIPOLIOMYELITIC HORSE SERUM CONCENTRATE AND ITS EXPERIMENTAL USE IN INFECTED MONKEYS

1. The horse, apparently itself unsusceptible to poliomyelitis, can be stimulated in certain cases but not all to the production of virucidal antibodies. 2. The virucidal potency of such immune serum can be raised to a point comparable to that of human convalescent serum and when concentrated and refined exhibits a four-fold increase in potency. 3. Such concentrates have proved effective in the prevention of paralysis in inoculated monkeys when given intraspinously before the onset of paralysis. 4. Treatment has been found more effective when therapeutic serums are given through the spinal route. 5. Pooled serum from "normal" adult donors has proved effective in neutralizing virus but its potency is approximately one-half that of convalescent serum.     

The heterogeneity of the serum activity of gamma-glutamyl transpeptidase in hepatobiliary diseases as studied by agarose gel electrophoresis.

Using electrophoresis on agarose gel, the serum activity of gamma-GT in various hepatobiliary diseases has been separated into several electrophoretic fractions. Predominance of activity was always found in fractions corresponding to either alpha1-, alpha2- or beta-globulins. The predominant activity fractions have been related to the various disease states and to the zymogram patterns of gamma-GT found in bile and tissue homogenates of liver, bile duct and pancreas. Bile disclosed activity in the beta-region, tissue homogenates of liver mostly in the alpha1- and alpha2- areas, bile dict showed very faint activity in the alpha1- position, while extracts of pancreas revealed activity in the regions of alpha2, alpha2-beta and beta-gamma. In serum, dominance of alpha1-activity was found in all cases associated with alcoholism and alcoholic cirrhosis, this activity also appeared as the most frequent dominant one in biliary obstruction. Predominance of alpha2 and/or the beta-fraction in most cases occurred associated with malignant processes involving liver and/or pancreas.     

Altered gamma-glutamyltransferase, aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase serum activities in long-term anticonvulsive therapy--comparison of diphenylhydantoin and carbamazepine

In 110 patients receiving a long-term anticonvulsant monotherapy with Diphenylhydantoin (DPH) and Carbamazepine (CBZ) the serum activities of gamma-GT, ASAT, ALAT, and AP were examined retrospectively. Elevated serum levels of enzymes were seen predominantly concerning gamma-GT and AP. 91% resp. 39% of patients receiving DPH-therapy showed increased gamma-GT resp. AP-levels compared to 64% and 14% of gamma-GT and AP-elevations by CBZ-treatment. All enzymes evaluated were more often and higher elevated by DPH than CBZ. Frequency and extent of increased activity of gamma-GT were highly related to daily dosage in both preparations. The proportion of pathological enzyme levels was associated with age in DPH and CBZ as well but not found to be significant. Sex differences in the frequency of increased enzyme activities could not be demonstrated. The results are discussed in the context of induction of cytochrome P-450-system.     

Serum gamma-glutamyl transpeptidase and physical exercise.

The immediate and delayed influence of exercise of variable type, duration and intensity on the serum gamma-glutamyl transpeptidase activity (gamma-GT) has been examined in athletes and untrained persons. The possible effects of exercise-induced changes in other parameters (plasma free glutamate, serum triglyceride, haemoconcentration) on the measured postexercise serum gamma-GT have been discussed, partly on the basis of our own experimental data. It appears that neither exercise itself, nor any one of the above mentioned factors (excepting a slight, transient effect of haemoconcentration after brief, intensive exertion) have noticeable influence on serum gamma-GT. Determinations of this enzyme for diagnostic purposes are therefore likely to be used without regard to acute and chronic exercise history in man.     

FURTHER OBSERVATIONS ON THE USE OF PNEUMOCOCCUS EXTRACTS IN EFFECTING TRANSFORMATION OF TYPE IN VITRO

Pneumococcus extracts highly active in inducing the in vitro transformation of the specific types of Pneumococcus have been prepared by dissolving S cells with sodium desoxycholate, precipitating the dissolved material in alcohol in which the bile salt remains soluble, and extracting the precipitate in salt solution. Further purification of these active extracts has been attained by the removal of considerable inactive material by charcoal adsorption and by reprecipitation of the adsorbed extract in alcohol or acetone. The importance of using young cultures for extraction, and of preventing autolysis during the preparation of the extracts, is emphasized. Extracts prepared by the method described have been filtered through Berkefeld Candles (V, N, and W) without appreciable loss in activity, provided the reaction of the extract was slightly alkaline at the time of filtration. The purified and filtered extracts are water-clear, and sterile by rigid cultural and animal tests. They have been heated to temperatures of 60°C. for 30 minutes without appreciable loss in their capacity to induce specific changes in type. And although they have generally shown definite decrease in potency after heating to temperatures above 80° C., some extracts have been found active even after an exposure of 10 minutes to a temperature of 90°C. They have been completely inactivated by boiling. Relatively small amounts of extract have been effective when added to a broth medium containing normal serum or serous fluid. In this medium, R pneumococci, irrespective of their type derivation, have developed and thereafter retained all the type-specific characteristics of the encapsulated S cells from which the extract was prepared. The specific action of the extracts is discussed with reference to their transforming and antigenic properties.     

Evaluation of a Dispensing Instrument (Dynatech MIC-2000) for Preparing Microtiter Antibiotic Plates and Testing Their Potency During Storage

The MIC-2000 96-channel dispenser was evaluated for accuracy and repeatability of dispensing 50-μl volumes of water. It was found to perform within the manufacturer's specifications for accuracy of ±5%, but not within the limits for repeatability of ±1%. Overall, instrument performance was found to be satisfactory for the antimicrobial susceptibility testing technique which has been implemented in this laboratory. A method is suggested for simple evaluation of the volumes dispensed. A variety of antibiotics may be stored in Trypticase soy broth in Microtiter plates without loss of potency for periods of at least 2 weeks at −20°C and 4 months at −70°C.     

STUDIES ON THE CULTIVATION OF THE VIRUS OF LYMPHOGRANULOMA VENEREUM

Two strains of the lymphogranuloma venereum virus were maintained in tissue cultures for 12 and 21 generations respectively. In a third strain a quantitative increase in potency as high as 1000 times was obtained by inoculating tissue cultures with infected emulsions of known virulence. Greater increases in potency were consistently obtained by maintaining tissue cultures and virus at room temperature (23°C. with a range of approximately ± 5°C.) than by incubating them at 37°C. The virus did not survive in the absence of oxygen. Embryonic guinea pig brain and serum ultrafiltrate were found to be the most effective vehicles for propagation of the lymphogranuloma virus. There is evidence that the site of activity for this virus is intracellular. Embryonic guinea pig brain cells were maintained in the serum ultrafiltrate diluted with buffered salt solution in good (morphologic) condition for as long as 70 days. Not only could old cultures be successfully inoculated with the virus of lymphogranuloma, but high titres could be maintained over extended periods.     

IMMUNOSUPPRESSION OF MICE INJECTED WITH HETEROLOGOUS ANTI-IMMUNOGLOBULIN HEAVY CHAIN ANTISERA

Neonatal injection of mice with rabbit anti-µ antiserum has been shown to produce complete loss of direct and indirect plaque-forming responses to sheep erythrocytes as well as loss of serum IgM and severe depressions of all other serum immunoglobulins. Similar injection of anti-γ1γ2 or anti-γ1 antibodies effects a loss of the indirect response but induces relatively minor alterations in serum Ig levels. Delaying initiation of anti-µ treatment until young adulthood results in a somewhat diminished effect on plaque-forming responses and serum Ig levels but triggers the release of high serum levels of an aberrant µ-bearing protein. Anti-µ suppression of genetically thymusless mice indicates that at least part of the target cells for suppression are bone marrow derived. A working hypothesis for the maturation of humoral antibody-producing cell lines as it relates to these data is discussed.     

EFFECTS OF MECHANICAL AGITATION AND OF TEMPERATURE UPON COMPLEMENT

1. Under certain conditions, mechanical agitation destroys the complementary activity of guinea pig serum. It is most injurious when carried out constantly at a temperature of 37° C., but it is extremely insignificant at 10° C. After the first few hours at 37° C., the destruction of complement proceeded much more rapidly, and after six hours it was almost complete. On the other hand, within one hour shaking had almost no destructive effect on complement, even at 37° C. From this we may conclude that the several shakings which are necessary for fixation experiments during incubation do not modify perceptibly the outcome of the reactions. 2. The rate of destruction of the complement of guinea pig serum at temperatures above 45° C. is progressively greater as it approaches 55° C., at which temperature the activity is reduced in thirty minutes to one-thirtieth to one-fortieth of the original strength of the unheated serum; but it is not completely destroyed, as is commonly assumed. The velocity of destruction of guinea pig complement when exposed to 55° C. for various lengths of time is found to be quite irregular, and not proportional to the length of time. This irregularity, however, presents a certain rhythm, a period of greater destruction alternating with one of less destruction.     

A Specific Inhibitor of Complement (C5)-Derived Chemotactic Activity in Serum from Patients with Systemic Lupus Erythematosus

In the course of examining polymorphonuclear leukocyte (PMN) chemotaxis in patients with systemic lupus erythematosus (SLE), we have found a previously undescribed serum inhibitor of complement (C5)-derived chemotactic activity. Serum from a 25-yr-old Black female with untreated SLE, when activated with zymosan, failed completely to attract either her own or normal PMN. Incubation of normal PMN with the patient's serum did not affect their subsequent random motility or chemotactic response toward normal zymosan-treated serum (ZTS). The patient's serum, however, did inhibit the chemotactic activity of normal ZTS and of column-purified C5-derived peptide(s), but had no effect on the chemotactic activity of either the synthetic peptide, N-formylmethionyl leucyl-phenylalanine or a filtrate prepared from a culture of Escherichia coli (bacterial chemotactic factor). The inhibitory activity in the patient's serum resisted heating at 56°C for 30 min and could be separated from C5-derived chemotactic activity in the patient's ZTS (or normal ZTS that had been incubated with the patient's serum) by chromatography on Sephadex G-75. Despite its effect on C5-derived chemotactic activity, the patient's serum did not influence two other C5-derived biologic activities: PMN lysosomal enzyme-releasing activity and PMN-aggregating activity. Chromatography of the patient's serum (65% ammonium sulfate pellet) on Sephadex G-200 yielded three distinct peaks of inhibitory activity. Two were heat labile and exhibited other properties of the previously described chemotactic factor inactivators of normal human serum. The third and most active peak, however, resisted heating at 56°C for 30 min, eluted with an apparent mol wt of 50,000-60,000, and acted specifically on C5-derived chemotactic activity. This uniquely specific, heat-stable inhibitor of C5-derived chemotactic activity has been found thus far in serum from 4 of 11 patients with active SLE and may account, in part, for altered host defenses against infections caused by pyogenic microorganisms.     

PREPARATION OF DRIED HEMOGLOBIN WITHOUT LOSS OF ACTIVITY

The technique for freezing, drying, and preserving in vacuo which is in common use for plasma can be successfully applied to hemoglobin solutions when the hemoglobin is first deoxygenated to the extent of 99.7 per cent or more. In confirmation of Morrison and Hisey, the preliminary deoxygenation of the solution is found necessary to avoid formation of methemoglobin during drying. If a solution of oxyhemoglobin is frozen and dried, 20 to 30 per cent is changed to methemoglobin. Deoxygnated hemoglobin dried and preserved in vacuo retained all its oxygenbinding activity for 180 days, when stored at temperatures from 4° to 30°C. Storage at 38°C. for 92 days, or at 56° for 7 days, caused no loss in activity. The dried hemoglobin had a foam structure which caused it to dissolve immediately upon contact with water. Deoxygnated hemoglobin in the dry state was partly converted to methemoglobin by even momentary contact with oxygen. When, however, the deoxygnated hemoglobin was dissolved before it was exposed to air, the hemoglobin in solution was relatively stable, and could be stored for months at 4° in contact with air without significant loss of activity.