Saline-soluble Proteins Research Articles

Evaluation of corn germ protein as an extender in plywood adhesive

This study was conducted to evaluate the potential of wet-milled corn germ protein as an extender in plywood adhesive. Partially defatted dried corn germ from wet-milling, containing 2.1% (dry basis, db) crude oil and 24.7% (db) crude protein, was ground to 40-mesh particle size to produce the meal. The predominant water- and saline-soluble proteins were extracted from the corn germ meal (CGM) by using 0.1 M NaCl as solvent in a method that used homogenization, centrifugation, dialysis, and freeze-drying. The recovered freeze-dried protein extract (FDPE) was substituted (on protein content basis) for wheat flour in the standard adhesive mix for sprayline coaters. Adhesive containing CGM was also prepared in the same manner. Mixing and adhesion properties of the corn germ-based adhesives were compared with those of the industry standard adhesive. The adhesive containing FDPE showed dispersibility and mixing behaviors, as well as viscosity values that were almost identical to those of the industry standard and superior to those of CGM adhesive. The mean wet tensile strength of the adhesive containing FDPE was notably greater (1.71 MPa) than that of CGM-based adhesive (1.34 MPa) and wheat-based standard adhesive (1.38 MPa), which indicated stronger bonding. The results demonstrated that corn germ protein has the potential to be an alternative protein extender in plywood adhesives for sprayline coaters.

Extensin network formation in Vitis vinifera callus cells is an essential and causal event in rapid and H(2)O(2)-induced reduction in primary cell wall hydration.

BackgroundExtensin deposition is considered important for the correct assembly and biophysical properties of primary cell walls, with consequences to plant resistance to pathogens, tissue morphology, cell adhesion and extension growth. However, evidence for a direct and causal role for the extensin network formation in changes to cell wall properties has been lacking.ResultsHydrogen peroxide treatment of grapevine (Vitis vinifera cv. Touriga) callus cell walls was seen to induce a marked reduction in their hydration and thickness. An analysis of matrix proteins demonstrated this occurs with the insolubilisation of an abundant protein, GvP1, which displays a primary structure and post-translational modifications typical of dicotyledon extensins. The hydration of callus cell walls free from saline-soluble proteins did not change in response to H2O2, but fully regained this capacity after addition of extensin-rich saline extracts. To assay the specific contribution of GvP1 cross-linking and other wall matrix proteins to the reduction in hydration, GvP1 levels in cell walls were manipulated in vitro by binding selected fractions of extracellular proteins and their effect on wall hydration during H2O2 incubation assayed.ConclusionsThis approach allowed us to conclude that a peroxidase-mediated formation of a covalently linked network of GvP1 is essential and causal in the reduction of grapevine callus wall hydration in response to H2O2. Importantly, this approach also indicated that extensin network effects on hydration was only partially irreversible and remained sensitive to changes in matrix charge. We discuss this mechanism and the importance of these changes to primary wall properties in the light of extensin distribution in dicotyledons.

Identification of GRASP-1 as a novel 97 kDa autoantigen localized to endosomes

We have identified an autoantigen that is recognized by antibodies from an 18-year-old female with a history of recurrent infections who later in her clinical course developed Raynaud's phenomenon and telangiectasias. By indirect immunofluorescence (IIF), the index serum produced a unique cytoplasmic discrete speckled (CDS) staining pattern that partially colocalized with early endosome antigen 1 (EEA1) but not Golgi complex or other cytoplasmic organelles in HEp-2 cells. When HEp-2 cells were treated with 0.1 N HCl, the cytoplasmic speckled staining of the index serum was markedly decreased, suggesting that the reactive antigen was soluble. Western blot analysis showed a reactive ∼97 kDa protein in a saline soluble protein preparation from HeLa cells. Mass spectrometric analysis of the excised 97 kDa band that was immunoprecipitated from HeLa cell extracts identified GRASP-1 as a possible target. The index serum and anti-GRASP-1 antibodies colocalized to structures in the cytoplasm of HEp-2 cells. Synthetic peptides representing the full-length GRASP-1 protein were used to identify reactive epitopes. Like many other cytoplasmic autoantigens, GRASP-1 has numerous coiled-coil domains throughout the protein with the exception of short segments at the amino and carboxyl terminus.

Non-Neoplastic Kidney Diseases, Abstract 110–113, Symposium

Renal antigens in mercuric chloride induced, anti-GBM autoantibody glomerular disease. Two antibody probes were used to characterize the putative renal antigens of HgCl2-induced antiglomerular basement membrane renal disease in Brown Norway (BN) rat. The first probe was the linear immunofluorescence imparting, in vivo bound, nephritogenic antiglomerular-basement-membrane autoantibody (anti-GBM-Ab). The second probe was a rat monoclonal antibody to the B subunit of laminin that was obtained from fusion of spleen cells of HgCl2 injected BN rat. By enzyme-linked immunosorbent assay (ELISA) the anti-GBM-Ab reacted with laminin, type IV collagen, collagenase-resistant noncollagenous portion of glomerular basement membrane (GBM), saline soluble proteins of kidney cortex homogenate and fibronectin. Western blot analysis of laminin indicated that the reactive epitopes detected by both probes were on the B chain subunit but not the A subunit. In nonreduced collagenase-digested GBM the epitopes were present on 27 kD and 42 to 48 kD polypeptides. A similar pattern was seen on collagenase-digested human GBM. On rat and human GBM the patterns obtained with rat autoantibody and autoantibody from a patient with Goodpasture syndrome were similar, suggesting that some of the in vivo bound anti-GBM autoantibodies in HgCl2-induced disease in rat are directed against epitopes which are similar to the Goodpasture antigen of human. Reactive epitopes were also detected on saline soluble proteins of kidney cortex homogenate with the predominant antigen being a 31 kD polypeptide. In the saline soluble proteins the reactive polypeptides including the major 31 kD polypeptide did not originate from laminin, type IV collagen, or the collagenase-resistant noncollagenous part of GBM. The precise structural origin of soluble proteins was not defined. The epitopes in type IV collagen and fibronectin could not be further characterized for molecular mass by Western blot analysis.

A cytoplasmic lectin produced by the fungus Arthrobotrys oligospora functions as a storage protein during saprophytic and parasitic growth.

It was recently shown that the nematode-infecting fungus Arthrobotrys oligospora contains a saline-soluble lectin (designated AOL) that is a member of a novel family of fungal lectins sharing similar primary sequences and binding specificities. During saprophytic growth in liquid cultures, levels of AOL and AOL mRNA were found to vary depending on the growth phase of the mycelium and the carbon/nitrogen (C/N) ratio of the medium. AOL was not detected in young mycelium. In older mycelium (stationary growth phase) grown in media with low C/N ratios (1 or 6), AOL comprised 5-20% of the total amount of saline-soluble proteins present in the mycelium. Neither the lectin nor its transcript was detected in mycelia grown in medium with higher C/N ratios (≥150). Under conditions of nitrogen starvation, AOL was preferentially degraded in relation to the total amount of saline-soluble proteins present in the mycelium. During the infection of nematodes, the level of AOL protein and AOL mRNA increased significantly once the nematodes had been penetrated and digested. Large amounts of AOL accumulated in the trophic hyphae growing inside the nematode as visualized by immunofluorescence microscopy. Later, AOL labelling was detected outside the digested nematodes, preferentially in strands of aggregated hyphae and in newly developed trap cells. Electron microscopy showed that AOL was localized to the cytoplasm and the nucleus of both vegetative mycelium and trap cells, and in the trophic hyphae growing inside the infected nematodes. These results indicate that AOL functions as a storage protein during both saprophytic and parasitic growth.

Kinetic Study and Modeling of the Degradation of the Maize Kernels Quality During Drying in Fluidized Bed

ABSTRACT Only a few kinetic studies have been reported in the literature on the evolution of comercial quality of maize during drying and to the best of our knowledge no model allows to predict the dynamic coupling of drying and product quality evolution. The aim of this work is to present new information on the effects of the operating conditions (harvest year, weight of maize to be dried, initial moisture content of the grain and air temperature) on the evolution of maize saline-soluble protein denaturation and the wet-milling quality during drying of maize in a batch floatation fluidized bed dryer. Also, kinetic laws are proposed for the quality criteria that will be combined with drying model already derived [1, 2]. The experimental results show that the degradation of the main components of maize (starch and proteins) is considerably affected by the temperature level and to a lesser extent by the initial moisture content of grains. Beyond 70°C, the denaturation of saline-soluble proteins occurs rapidly in the heat-up period of the grains. As for the wet-milling quality degradation, it starts only above 90°C. Kinetic laws derived from this study express the variation of the degradation rate of proteic and wet milling quality as functions of the solubility of saline-soluble proteins or the starch-gluten separation index, the grain moisture content and temperature.

THERMAL EXTRUSION and ALKALI PROCESSING of DRY BEANS (PHASEOLUS VULGARIS L.)

Dry beans are an important source of protein but contain antinutritional factors. an important antinutrient is phytohemagglutinin (PHA), a heat-labile lectin known to depress the nutritional quality of dry beans. This work was undertaken to evaluate the utility of two processes that may aid in the inactivation of PHA, extrusion and cooking in a high pH medium. Extrusion was found to be relatively ineffective in reducing the activity of PHA of whole red kidney or black beans. Extrusion was more effective but highly variable in reducing the PHA activity of bean flours (25-80%). However, soaking and cooking beans at high pH was very effective, significantly reducing the activity of PHA and also reducing the time required to reach a palatable texture. Soaking and cooking dry beans at high pH also caused significant changes in the saline soluble protein extract as determined by gel electrophoresis. High pH cooking treatment may be useful in improving nutritional quality of dry beans.

The effect of adjuvant and specific or non-specific vaccination on development of protective immunity of rabbits against Trichostrongylus colubriformis infection

Adult New Zealand rabbits were vaccinated subcutaneously with one dose of 100 μg adult nematode phosphate buffered salinesoluble proteins (PBS-ASP, groups I and II), a detergent-soluble fraction of adult somatic proteins (DS-ASP, group III) or three doses of l mg normal rabbit serum proteins (group IV). Injections of the immunogens in groups II, III and IV were accompanied with beryllium hydroxide, Be(OH) 2 as an adjuvant. Vaccinated rabbits and also those of group V (naive) were challenged orally with 10,000 infective larvae of T. colubriformis 14 days after antigen injection and necropsied 2 weeks later. A single dose of PBS-ASP induced 33.5% protection when the antigen was given alone (group I) and 69.4% when injected with Be(OH) 2 (group II). A detergent-soluble fraction of ASP given with the adjuvant provided 87.2% protection (group III), whilst non-specific vaccination with serum proteins plus Be(OH) 2 elicited 99% protection (group IV). Mesenteric lymph node leukocyte responses were measured using a leukocyte migration inhibition assay. A significant response was observed only in group IV. In ELISA tests IgA antibodies specific to PBS-ASP reached the highest level in the intestinal mucosa of groups I and II and in the bile of groups I and III. Antibody levels of IgG isotype were similar in the intestinal mucosa of all the immunized groups. Nematode antigen was detected using a ‘sandwich’ ELISA method in faecal protein extracts of rabbits of groups II and III on days 2–6 after challenge.

Solubility of amaranth seed proteins in sodium sulphate and sodium chloride: the main factor in quantitative extraction for analysis

Summary.Nitrogen was extracted more efficiently from amaranth seed with 0.04 M Na2SO4 (5% w/v) than with either 0.09 M or 0.17 M NaCl (5% or 10% w/v), despite both solutions having the same ionic strength (μ= 1). Solubility of saline soluble proteins (albumin ± globulin) was very poor in either water or 1M NaCl, but increased in 0.4M NaCl at alkaline pH between 7 and 10. Globulins were very soluble in 0.4M NaCl at a pH 9. Albumin was the main storage protein. Saline soluble proteins formed very weak gels.

A soybean cell wall protein is affected by seed color genotype.

The dominant I gene inhibits accumulation of anthocyanin pigments in epidermal cells of the soybean seed coat. We compared saline-soluble proteins extracted from developing seed coats and identified a 35-kilodalton protein that was abundant in Richland (genotype I/I, yellow) and much reduced in an isogenic mutant line T157 (genotype i/i, imperfect black seed coats). We purified the 35-kilodalton protein by a novel procedure using chromatography on insoluble polyvinylpolypyrrolidone. The 35-kilodalton protein was composed primarily of proline, hydroxyproline, valine, tyrosine, and lysine. Three criteria (N-terminal amino acid sequence, amino acid composition, and sequence of a cDNA) proved that the seed coat 35-kilodalton protein was PRP1, a member of a proline-rich gene family expressed in hypocotyls and other soybean tissues. The levels of soluble PRP1 polypeptides and PRP1 mRNA were reduced in young seed coats with the recessive i/i genotype. These data demonstrated an unexpected and novel correlation between an anthocyanin gene and the quantitative levels of a specific, developmentally regulated cell wall protein. In contrast, PRP2, a closely related cell wall protein, was synthesized later in seed coat development and was not affected by the genotype of the I locus.

Nucleotide Sequence of a cDNA Clone Corresponding to the Maize Globulin-2 Gene

Maize globulins represent a group of saline-soluble proteins that accumulate primarily in the developing embryo (4, 6, 7). The most abundant globulin components are the Mr 63,000 GLB1 protein and the Mr 45,000 GLB2 protein, which are encoded by the Globulin-i (Glbl) and Globulin-2 (Glb2) genes, respectively (4, 7). GLB 1 and GLB2 collectively comprise 10 to 20% of the protein present in the mature maize embryo. The amino acid composition and the developmental accumulation pattern of GLB 1 and GLB2 suggest that these are embryo storage proteins (4). GLB1 exhibits sequence similarity to 7S seed globulins of both monocots and dicots (2, 3). A common characteristic of seed storage proteins described to date is that they are encoded by multigene families (8). In contrast, the Glbl gene exists as a single copy in the haploid genome as determined by genetic analyses of several electrophoretic variants (7) and as recently confirmed at the molecular level by gene copy-number reconstruction analysis (2). Although we have not observed electrophoretic variants for GLB2, two null alleles for Glb2 have been identified (Table I). GLB2 also appears to be encoded by a single gene as determined by genetic analyses in which each of the Glb2 null alleles segregates as a Mendelian recessive trait and by Southern blot analysis of maize DNA in which Glb2-specific clones are used as probes. The availability of allelic variants for both of the single-copy maize Glb genes allows for detailed analysis of allelic polymorphism in plants at the molecular level. Recently, we reported (2) the isolation and characterization of a cDNA clone corresponding to the Glb]-S allele, which represents the first monocot 7S globulin to be analyzed at the molecular level. We present here the nucleotide sequence of a full-length cDNA clone corresponding to Glb2. The amino acid sequence deduced from the pcGlb2 nucleotide sequence (Fig. 1) indicates that GLB2 is synthesized as a preproprotein, as is the case for GLB 1 (2, 5) and other seed storage globulins (8). Alignment of mature GLB 1 and GLB2 from their amino termini indicates that the two polypeptides share 28% amino acid sequence identity. Although a slight amount of immunological cross-reactivity has been observed for the GLB2 and GLB1 polypeptides, the two cDNA clones pcGlb2 and pcGlb1S do not cross-hybridize at a Tm of -25?C. Efforts are underway to isolate genomic clones corresponding to Glb2 as well as the null alleles Glb2-0 and Glb2-NJ to determine the molecular basis of the null phenotype in each case.

Evaluation of a Sound Velocity Analyzer for Estimating Maximum Internal Temperature to Which Meat Products Have Been Heat Processed

The sound velocity (SV) values of a variety of solutions (distilled water, saline, silicon dioxide, bovine serum albumin, coagulated beef protein) were determined as the temperature of the solution was increased from 10 or 20° to 71°C. Few differences were found in the SV values for the various types of distilled water; however, there was an increase in SV values as the concentration of saline increased. Level of silicon dioxide or bovine serum albumin had no significant effect on SV values; however, there was an increase in SV values as percentage of precoagulated beef protein solids in solution increased. The sound velocity analyzer used in this study was found not to be sensitive enough to detect the onset of coagulation of saline soluble proteins and, therefore, would not lend itself as an objective method which could be related to end-point temperature of meat products.

Renal antigens in mercuric chloride induced, anti-GBM autoantibody glomerular disease

Two antibody probes were used to characterize the putative renal antigens of HgCl2-induced antiglomerular basement membrane renal disease in Brown Norway (BN) rat. The first probe was the linear immunofluorescence imparting, in vivo bound, nephritogenic antiglomerular-basement-membrane autoantibody (anti-GBM-Ab). The second probe was a rat monoclonal antibody to the B subunit of laminin that was obtained from fusion of spleen cells of HgCl2 injected BN rat. By enzyme-linked immunosorbent assay (ELISA) the anti-GBM-Ab reacted with laminin, type IV collagen, collagenase-resistant noncollagenous portion of glomerular basement membrane (GBM), saline soluble proteins of kidney cortex homogenate and fibronectin. Western blot analysis of laminin indicated that the reactive epitopes detected by both probes were on the B chain subunit but not the A subunit. In nonreduced collagenase-digested GBM the epitopes were present on 27 kD and 42 to 48 kD polypeptides. A similar pattern was seen on collagenase-digested human GBM. On rat and human GBM the patterns obtained with rat autoantibody and autoantibody from a patient with Goodpasture syndrome were similar, suggesting that some of the in vivo bound anti-GBM autoantibodies in HgCl2-induced disease in rat are directed against epitopes which are similar to the Goodpasture antigen of human. Reactive epitopes were also detected on saline soluble proteins of kidney cortex homogenate with the predominant antigen being a 31 kD polypeptide. In the saline soluble proteins the reactive polypeptides including the major 31 kD polypeptide did not originate from laminin, type IV collagen, or the collagenase-resistant noncollagenous part of GBM. The precise structural origin of soluble proteins was not defined.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative HPLC analysis of human plaque proteins in coronary and thoracic aorta arteries

Quantitative HPLC analysis of saline-soluble proteins obtained from human coronary and thoracic aorta plaque and from whole internal mammary artery were performed. Protein extracts were characterized by anion exchange and reverse-phase HPLC and the integrated chromatographs revealed significant differences in both peak retention times and areas for protein species from coronary artery compared to thoracic aorta artery plaque. Coronary artery plaque proteins possessed a high degree of cationic charge and polarity compared to those present in thoracic aorta plaque and normal mammary artery. This suggests that specific protein markers may be expressed in plaque of different anatomical origin, and that the processing of protein may be distinct to plaque sites. In contrast, characterization of molecular weight by gel electrophoresis resolved no major differences between plaque types. These findings indicate that proteins in human plaque lesions of different anatomical origin can be resolved by HPLC methodology and that they exhibit different charge and polarity. Such an HPLC approach may prove useful in the quantitative identification and ultimate isolation of specific protein markers present in plaque during atherogenesis, and in the study of mechanisms of protein involvement in plaque formation.

Modulation of Differentiation by Retinoids

At high levels of vitamin A and other retinoids (3 X 10(-6) M) attachment of human keratinocytes to 3T3-coated plastic dishes is mildly inhibited. Retinoids at this concentration in culture media seem to have an antikeratinizing effect in that cells appear to be less differentiated. Retinoid-treated cultures are less stratified, having fewer cell layers, and display larger intercellular spaces and rounder, less flattened cells. Treated cultures also contain higher percentages of saline-soluble proteins and lower percentages of proteins requiring reduction and/or denaturants for solubility. This suggests that in treated cultures, the most keratinized cells are absent. Growth curves show that those most keratinized cells are sloughed from the dish and appear in the media. Thus at 3 X 10(-6) M, the major retinoid effect is to promote desquamation. At higher concentrations, retinoids are toxic to the keratinocyte, but at lower concentrations, they may be stimulative.

Differential Electrophoretic Behavior Among Strains of Aedes Aegypti (Diptera: Culicidae)

Numerical analysis of the mobilities of salinesoluble proteins revealed differential patterns of electrophoretic behavior among 3 strains of Aedes aegypti . Females exhibited much greater contrast and divergence in their electrophoretic patterns than did males. The electrophoretic variability demonstrated in this study is considered with respect to its possible physiological and genetic basis.

Electrophoretic and histochemical comparison of three strains of Aedes aegypti

1. 1. Saline-soluble proteins of young adults of 3 strains of the mosquito, Aedes aegypti, were compared by means of electrophoretic and histochemical techniques. 2. 2. Several electrophoretic fractions were found to be histochemically complex, possessing conjugated lipid and carbohydrate moieties as well as esterase and phenol oxidase activities. 3. 3. Although the distribution of proteins and esterases generally followed strain-specific patterns, the sexes of each strain were electrophoretically and histochemically distinguishable.

Hemodynamically-induced increase in soluble collagen in the anastomosed veins of experimental arteriovenous fistulae

External jugular veins that had been subjected to the hemodynamic stresses produced by experimental arteriovenous anastomosis developed 2% increased total protein contents and 17% increased collagen contents. When the stressed veins were homogenized and extracted with saline solutions, statistically significant increases in the saline-soluble proteins and in the saline-soluble collagen (87% and 267%, respectively) were observed. Increased amounts of low molecular weight peptides were found in the extracts. A fraction of these peptides could be degraded by Clostridium collagenase. The saline extract also contained proteins which resembled by their amino-acid composition the acidic structural proteins of the connective tissues. Additonally, in 3 dogs so tested, changes were found in the hydroxylation and glycosylation of lysine from gelatin extracts as well as in the lysine and desmosine contents of the insoluble elastin fractions. This is the first demonstration of a hemodynamically induced increase in the saline solubility of connective tissue proteins in the absence of dietary manipulations.

Chemosensory Mechanism in American Cockroach Olfaction and Gustation

ALTHOUGH numerous theories of olfaction and gustation1–8 have been published, there is little experimental evidence on the energy-transfer mechanisms involved in the interactions of chemical messengers with proteinaceous receptors of sensory neurones. One such mechanism, involved in the inhibition of feeding of Periplaneta americana and Scolytus multistriatus by various 1,4-naphthoquinones, has been extensively studied and partially interpreted behaviourally9–16, ultrastructurally10–16, electrophysiologically10–16 and biochemically10–16. These messenger naphthoquinones were shown to act through a selective reaction with sulphydryl10–16 groups in a particular membrane-associated receptor macromolecule which was proven to be physically accessible in situ17 to these chemicals through cuticular pores and tubules10–16 in the walls of chemoreceptor sensillae on the insect's antennae. This specific energy transfer resulted in disulphide-bond formation or thiol complexing10–16, both of which are proven18 mechanisms in certain proteins for causing conformational changes. Such changes can alter the cationic permeability of macromolecules composing membranes18. The messenger-altered cation permeability of the receptor thus can result in the generation of action potentials in the sensory neurone10–16. Here we report certain physicochemical characteristics of the messenger-naphthoquinone receptor protein interactions as analysed in a deoxygenated system at physiological pH by the in vitro technique of dropping mercury electrode (DME) polarography (PO4 ‘Polariter’, London Co.). The polarogram is a current–voltage curve obtained by applying a gradually increasing voltage to a system and measuring the current flow at each potential. Characteristic polarographic half-wave potential (that is, E ½) values for proteins were attributed exclusively to exposed, reactive sulphydry1, or reducible disulphide, groups19–22. In our own investigations, the ‘Triton’ X-100-solubilized quinone-reactive receptor protein from the cockroach antennae gave a characteristic E ½ of −1,860 mV. A control (non-quinone receptor) protein system composed of the saline-soluble proteins from the same antennae gave an E ½ of −1,800 mV. Adding messenger 5-hydroxy-1,4-naphthoquinone (5 × 10−5M) in the receptor preparation characteristically shifted the E ½ from −1,860 to −1,880 mV; another less repellent, or feeding-inhibitory, messenger, 2-methyl-1,4-naphthoquinone (5 × 10−5 M), only shifted the receptor E ½ to −1,875 mV. Neither of these messenger quinones altered the −1,800 mV E ½ of the non-receptor control protein. N-Ethyl maleimide (NEM), a highly specific thiol reagent, at 5 × 10−5 M in the receptor preparation shifted the E ½from −1,860 to −1,890 mV. In this −1,890 mV state, the receptor was non-reactive with messenger quinones. Treatment of the receptor in the −1,890 mV state with the disulphide reducing agent, dithiothreitol (DTT), brought the E ½ of the receptor back down to −1,860 mV. In this restored −1,860 mV state, the receptor again reacted with messenger quinones.

Immunosuppressive alpha globulin from bovine thymus: I. Preparation and assay

DEAE chromatography at pH 5.0 of the saline-soluble proteins from bovine thymus glands yields a protein fraction similar in activity to the immunosuppressive alpha-2 globulins previously described from bovine and human serum. Of 32 preparations 16 had consistent and reproducible suppressive activity to DNA synthesis in phytohemagglutinin (PHA)—stimulated lymphocytes in concentrations ranging from 50–600 μg/ml in vitro. In vivo immunosuppression, not directly related to the degree of in vitrolymphocyte suppression, occurred in doses of 4–5 mg per mouse, and was assessed by the hemagglutinin response to sheep red blood cells. A variety of other protein and nucleic acid fractions were not suppressive in these assays; in particular, fractions A and B, which precede the immunosuppressive Fraction C in the elution from DEAE-cellulose, are neither suppressive, nor do they significantly alter the effects of Fraction C.