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[Analysis of lipoproteins using polyacrylamide gel electrophoresis and fractionated precipitation with polyanions and detergents. Use in the classification of hyperlipoproteinemias].

Sodium dodecylsulphate precipitates very low density lipoproteins (VLDL and chylomicrons) rich in triglyceride and a narrow correlation (r = 0.9) was found between the turbidimetric index SDS and triglyceridemia. Heparin in calcium medium acts in the same way and precipitates also low density lipoproteins (LDL) rich in cholesterol. A correlation (r = 0.875) was drawn up between the cholesterol LDL and the difference between the turbidimetric indices (heparin Ca -- SDS). In the first case, we were thus measuring by turbidimetry a VLDL + chylomicron index and, in the second case, an LDL index which permits one to obtain simplified typing of the hyperlipoproteinemias. A nomogram linking the two indices of hyperlipoproteinemia type was drawn up experimentally. This orientation analysis may be completed by electrophoresis on polyacrylamide gel used in concentration gradient and pH. In the system proposed, the lipoproteins were previously stained with tetrazolium nitroblue and clearly shown up. The chylomicrons in particular, become separated from the VLDL, the sinking pre-beta-lipoprotein or Lp (a) was identifiable and the type III hyperlipemia was easily diagnosed.

Adolescent↗

Precipitation test--a rapid method for diagnosis of tuberculous meningitis.

Precipitation test (modified Lavinson's test) was conducted in 125 cases of meningitis. Of these, 50 cases (proved cases of tuberculous meningitis) who served as positive controls showed positive response for the precipitation test in 96% and in 25 cases of negative control group (cases of meningitis other than TBM), it showed only 4% positivity. The remaining 50 cases of TBM where the cytological and biochemical findings were not in correlation with clinical diagnosis of TBM, showed 92% positivity for early diagnosis of TBM.

Benzenesulfonates↗

Dextran sulfate precipitable (light) plasma lipoproteins in genetically active and passive rats fed two types of diet.

Genetically active and passive female rats were fed a purified sucrose/fat diet or a stock diet from the age of 11 to 27 weeks. The age-related increase in plasma total cholesterol concentration was much more pronounced in active rats fed the purified diet than in active rats fed the stock diet. No significant alteration with age took place in the plasma cholesterol level of passive rats fed the stock or the purified diet. The increase in body weight was somewhat greater in active and passive rats fed the purified diet than in those fed stock diet. The lipoprotein distribution at the age of 27 weeks was estimated using both polyacrylamide gel electrophoresis and dextran sulfate/Ca++ precipitation. As compared with the stock diet, the purified diet effected a decrease in the relative amount of prebeta and beta lipoproteins in both active and passive rats. The major effect was, however, a pronounced increase in the relative amount of dextran sulfate precipitable "light prealpha" lipoproteins with no effect on "heavy prealpha" or alpha lipoprotein percentages. Alpha lipoprotein percentages were slightly higher in active than in passive rats. The results confirm our earlier observation that there might be a dissociation between the dietary effect on the plasma total cholesterol level and on the lipoprotein distribution. The data also suggest that the hyperlipemic sucrose/fat diet causes a shift from the prebeta and beta lipoproteins to the "light prealpha" fractions.

Animals↗

Comparison of improved precipitation methods for quantification of high-density lipoprotein cholesterol.

We compared the standard Lipid Research Clinics heparin-Mn2+ (46 mmol/L) method and five improved precipitation methods for quantification of high-density lipoprotein (HDL) cholesterol. Three of these methods--a dextran sulfate-Mg2+ procedure, reported as a Selected Method, a modified heparin-Mn2+ (92 mmol/L) method, and a modified phosphotungstate-Mg2+ procedure--all gave similar results. Three other methods--the standard heparin-Mn2+ (46 mmol/L) method and two polyethylene glycol methods (75 g/L or pH 10 reagent at 100 g/L final concentrations)--gave slightly higher values for HDL cholesterol. Addition of NaCl or glucose to specimens did not significantly change protein precipitation. In terms of sedimentation effectiveness with hypertriglyceridemic specimens, the methods were ranked in the following order: polyethylene glycol (pH 10, 100 g/L) greater than dextran sulfate-Mg2+ greater than heparin-Mn2+ (92 mmol/L) = polyethylene glycol (75 g/L) greater than phosphotungstate-Mg2+ greater than heparin-Mn2+ (46 mmol/L).

Adolescent↗

Thermodynamics of water-induced precipitation of cholesterol and its acetate, benzoate and stearate derivatives dissolved in 1,4-dioxane and 2-propanol.

The energetics of the precipitation process depended on the solute-solvent combination and the enthalpy and entropy of precipitation compensated each other. The partial molal volumes of the lipids in both the solvents were greater than the anhydrous molar volumes, except for cholesterol in 1,4-dioxane and cholesteryl acetate in 2-propanol where the order was reverse. While the partial molal compressibilities of all the solutes studied were negative in 1,4-dioxane, those of cholesterol and cholesteryl acetate in 2-propanol were, respectively, negative and positive. The negative values were supported by considerable solvation of the solutes, particularly in 1,4-dioxane.

1-Propanol↗

[Clinical effectiveness of the method of extracorporeal heparin precipitation of plasma proteins (selective plasmapheresis) in patients with immune complex pathology].

The authors analyse the efficacy of extracorporeal heparin precipitation of plasma proteins (selective plasmapheresis) in patients with immune complex pathology. The method lies in extracorporeal precipitation at +4 degrees C of fibronectin, cryofibrinogen and other macromolecular complexes with the aid of heparin, followed by plasma return to the patient. The method was applied to 39 patients. Hemorrhagic microthrombovasculitis was diagnosed in 7, vasculitis paralleled by hepatosplenomegaly of unclear genesis in 1, generalized vasculitis with marked systemic manifestations in 1, immune complex vasculitis complicated by recurrent polychondritis in 1, chronic active hepatitis associated with immune complex vasculitis in 1, rheumatoid arthritis in 8, systemic lupus erythematosus in 4, spleen lymphoma and Sjögren's syndrome in 1, idiopathic amyloidosis in 2, multiple sclerosis in 10 patients. Altogether 309 sessions of selective plasmaphapheresis were carried out. The time course of the clinical and laboratory criteria of the disease activity were studied before and after each plasmapheresis session. It was ascertained that the method is highly effective in the treatment of hemorrhagic vasculitis. It is mostly used if conventional therapy with heparin, cryoplasm, and deaggregating agents appears abortive. Apparently, it is the only method that may rapidly arrest the abdominal syndrome associated with hemorrhagic vasculitis. The possibility and advisability of the use of chronic plasmapheresis are discussed. Application of the method to the treatment of rheumatoid arthritis permits reducing the dose of steroid hormones, thereby not stimulating the disease activity. The method is the therapy of choice in different diseases associated with polyvalent allergy. In amyloidosis and multiple sclerosis, the method efficacy is questionable.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

Free and total insulin as determined after precipitation with polyethylene glycol: analytical characteristics and effects of sample handling and storage.

We evaluated results of radioimmunoassays of free and total insulin after precipitation of endogenous antibodies with polyethylene glycol (PEG), and we investigated the influence of collection time, temperature, and storage in heparin- cr EDTA-treated plasma or serum on results for free insulin. Analytical recovery of free insulin was 99.3%, of total insulin 96.4%. For free insulin, assay precision (CV) was 4.0-13.0% (intra-assay) and 7.8-10.7% (inter-assay); for total insulin, 3.6-9.5% and 6.6-11.7%, respectively. Free insulin decreased in plasma (p less than 0.05) and serum (p less than 0.01) at room temperature after 3 h and in promptly analyzed serum (p less than 0.01). Storage of samples at -20 degrees C increased the concentration of free insulin in plasma (p less than 0.025) and serum (p less than 0.005), whereas the free insulin content of supernates after PEG precipitation was stable, except for a slight decrease in serum samples (p less than 0.02). We conclude that, for radioimmunoassay of free and total insulin, plasma should be used, treated with PEG without delay; supernates then are analytically stable for as long as 26 weeks at -20 degrees C.

Antibodies↗

Precipitation of serum proteins by the lectin from wheat-germ.

We investigated the composition of the precipitate that forms when wheat-germ lectin derived from Triticum vulgaris is added to serum. A number of serum proteins are precipitated, representing about 2.5% of the total serum protein. This study demonstrates that the interaction of this lectin with the bone isoenzyme of alkaline phosphatase is not specific.

Alkaline Phosphatase↗

[Cholesterol, phospholipid and apoB composition of LDL: comparison of precipitation and ultracentrifugation methods].

Cholesterol, phospholipid and apoB levels were determined in LDL precipitated by amphiphatic polymers (Biomérieux kit, Marcy-l'Etoile, France). Intra-assay and inter-assay analysis performed on 4 serum pools were satisfactory. For 113 sera (triglyceride level less than 4.5 mmol/l, absence of VLDL-remnants), the 3 variables were well correlated with those of centrifuged LDL (0.93 less than r less than 0.98); however, LDL-cholesterol values were significantly higher (p less than 0.001), and those of LDL-phospholipids (p less than 0.001) and LDL-apoB (p less than 0.005) significantly lower. The correlations between the 3 constituents studied gave statistically different regression coefficients for the two methods. It is concluded that these precipitated lipoproteins do not seem to correspond to LDL but to "LDL markers".

Apolipoproteins B↗

Anti-IgG combined with rate nephelometry for measuring polyethylene glycol-precipitated circulating immune complexes.

Immune complexes from serum were assayed for IgG by a simple rate-nephelometric method after extraction with polyethylene glycol that removes monomeric IgG. A 30-min preincubation of the extracted material in reaction buffer before the anti-IgG is introduced eliminates falsely increased values owing to precipitation that increased baseline light scatter in the reaction buffer. We found good parallelism in the reaction of anti-IgG with the IgG calibrator, aggregated human globulin, or endogenous immune complex. Thus IgG can be used for calibration in place of aggregated human globulin, greatly simplifying the assay. A good correlation was found between the present assay and the C1q-binding test (r = 0.83). The present assay is both sensitive and reproducible. The extraction and assay are straightforward and can be completed in a single morning after an overnight precipitation. The reagents for extraction are easily prepared and inexpensive, and the materials for assay are available in kit form. We believe this approach to be well suited for many clinical laboratories to measure circulating immune complexes.

Antigen-Antibody Complex↗

Measurement of neonatal equine immunoglobulins for assessment of colostral immunoglobulin transfer: comparison of single radial immunodiffusion with the zinc sulfate turbidity test, serum electrophoresis, refractometry for total serum protein, and the sodium sulfite precipitation test.

Four procedures for assessment of adequacy of colostral immunoglobulin (Ig) transfer in foals were evaluated. Results of zinc sulfate turbidity test, serum electrophoresis, total serum protein refractometry, and sodium sulfite precipitation test were compared with immunoglobulin G content determined by single radial immunodiffusion. The zinc sulfate turbidity test gave acceptable results for IgG, except that hemolyzed serum samples gave higher than expected values. A correction factor for hemolyzed serum was found to be useful. Serum electrophoresis was a satisfactory method of estimating IgG content. Total serum protein values may not be a valid basis for estimating IgG content, inasmuch as postsuckling total protein values were found to decrease in some foals in which passive transfer of IgG had been adequate. Sodium sulfite precipitation reactions were too unpredictable to be of value for determination of neonatal IgG concentration.

Animals↗

[Occurrence and behavior of fibrin fibrinogen degradation products in concentrates of factor VIII, cryoprecipitate and dextran precipitate during long-term preservation].

In deep-frozen and lyophilized cryoprecipitates and dextran precipitates it was analyzed by the TRCHII and the SCT, whether these factor VIII concentrates contain fibrinogen fission products effecting as inhibitors of the coagulation of blood and how they act during storage for 200 days. Simultaneously the parameters total protein, fibrinogen, fibrin monomers, plasminogen and factor VIII activity were always determined. In all preparations being tested fibrinogen fission products could be detected. Lyophilized cryoprecipitates showed the highest average values with 7.97 microgram/ml (TRCHII) resp. 18.31 microgram/ml (SCT). They were followed by deep-frozen cryoprecipitates with 6.93 microgram/ml (TRCHII) resp. 7.54 microgram/ml (SCT). Deep-frozen dextran precipitates free of heparin contained the fewest fission products (TRCHII: 0.36 microgram/ml; SCT: 3.13 microgram/ml). At the beginning of the storage only few fission products were found in all groups of preparates. A predominantly clear rise with forming a peak after about 115 to 145 days and a drop following happened with extended storage period. This course of storage of the quantities of fission products shows that the fibrinogen fission products arise during storage.

Blood Preservation↗

Cellulolytic activity of moulds. II. Various methods of precipitating and concentrating enzymes and their influence on the activity of cellulolytic preparation and xylanase of Aspergillus terreus F-413.

Complexes of cellulolytic enzymes and xylanase were precipitated and concentrated by various methods from post-culture liquids of Aspergillus terreus F-143, containing cellucotton as carbon source. The best results in regard to the specific activity of the preparations were obtained by precipitation of enzymes with acetone-denatured ethanol. Besides high cellulolytic and xylanase activity the crude enzyme preparation showed the presence of small amounts of amylase, protease and polygalacturonase.

Acetone↗

Adjuvant effect of bacterial LPS and/or alum precipitation in responses to polysaccharide and protein antigens.

A conjugate of a hapten (NIP) and a strongly antigenic protein chicken gamma globulin (CGG), when injected in soluble form into mice, induced weak primary responses, as weak as responses induced by conjugates of NIP (or other haptens) to polysaccharides Ficoll or alpha (-1-6) dextran. Mean concentrations of anti-hapten antibodies on day 14 varied within the range of 37-105 micrograms/ml (C57BL mice) or 14-38 micrograms/ml (CBA mice). The NIP-protein conjugate administered in alum-precipitated form induced 100 times higher primary antibody responses. Alum-precipitation of NIP-Ficoll made it a modestly stronger antigen than soluble NIP-Ficoll. When lipopolysaccharide (LPS) was injected together with any of the soluble antigens, the mice produced plenty of anti-hapten antibodies regardless of whether the antigen was hapten-polysaccharide or hapten-protein conjugate. Concentrations on day 14 varied from around 400 micrograms/ml to approximately 1600 micrograms/ml. LPS had a similar adjuvant effect on antidextran responses. LPS alone induced a polyclonal immunoglobulin production, and the immunoglobulin produced included 'anti-NIP' or 'anti-dextran' detectable in the solid phase antibody assay. These 'antibodies' induced by LPS alone were almost totally mercapto-ethanol-sensitive and poorly detectable by Farr assay or the bacteriophage assay. The response to the LPS+antigen combination was specific for the antigen and included both mercapto-ethanol-sensitive and resistant antibodies.

Adjuvants, Immunologic↗

A comparison of an australian bluetongue virus isolate (CSIRO 19) with other bluetongue virus serotypes by cross-hybridization and cross-immune precipitation.

No major differences in size were observed when both the double-stranded RNA and the polypeptides of the Australian bluetongue virus (BTV) isolate CSIRO 19 (BTV-20) were compared with those of other BTV serotypes such as BTV-10 and BTV-4. Minor capsid polypeptide P6 of both BTV-20 and BTV-4, which electrophoreses as a single band on continuous phosphate buffered gels, in separated into 2 distinct bands on discontinuous glycine-buffered gels. This was not the case with BTV-10. Cross-immune precipitation of BTV-20 with BTV-10, BTV-17, BTV-4 and BTV-3 indicated strong immunological cross-reaction of the group-specific antigen P7 of the different serotypes. There was also some cross-immune precipitation of the serotype-specific polypeptide P2 of BTV-20 and BTV-4. This result is in agreement with the observed cross neutralization of these 2 viruses. The main distinction between BTV-20 and the other BTV serotypes was observed in cross-hybridization experiments. The homology between the nucleic acid of BTV-20 and other BTV serotypes was less than 30%, whereas homology normally found between BTV serotypes is at least 70%. The hybridization products of the different BTV serotypes were analysed by electrophoresis and fluorography. Two main hybrid segments were observed in all heterologous hybridizations with BTV-20 as a compared with 7 hybrid segments in hybridizations between BTV-4 and BTV-10. In order to determine from which genome segment of BTV-20 these 2 hybrid segments were derived, the hybridizations were carried out with individually purified double-stranded RNA segments. These results indicate that the 2 segments of BTV-20 that show the largest homology to corresponding segments of a heterologous BTV serotype are No. 7 and 10.

Bluetongue virus↗

Two cellular proteins that are precipitated by an antitumor-RSV rat serum.

Here we report studies on non-viral proteins specific for transformation precipitated by immune sera. These sera were from rats injected by PR-RSV transformed rat XC cells and from rabbits immunized by subcutaneous injections of cytoplasmic membranes from rat cells transformed by Fujinami avian sarcoma virus. Immune complexes were analyzed in one- or two-dimensional gel electrophoresis and by autoradiography. Only by using antisera obtained from rats bearing XC-cells induced tumors, two proteins were identified with an apparent molecular weight (Mr) of 105 000 (p105) and 98 000 (p98) and an isoelectric point of 6.4 and 6.2, respectively, in extracts of mammalian cells transformed by avian sarcoma viruses (ASV). These proteins were common to all ASV-transformed cell lines. In kinase reaction, the protein p98 can be phosphorylated. Similar proteins were neither detected in normal nor in chemically transformed cells. None of the proteins were precipitated with control serum from rat, rabbit, and goat.

Animals↗

Analysis for nickel in plasma and urine by electrothermal atomic absorption spectrometry, with sample preparation by protein precipitation.

We describe and evaluate a method for determining nickel in plasma and urine by atomic absorption spectrometry. Proteins are precipitated with trichloroacetic acid and sulfuric acid; ammonium pyrrolidinedithlocarbamate is used as the chelating agent for nickel, and methyl isobutyl ketone as extraction solvent. The results were compared with results obtained by the acid-digestion technique for removing proteins and other organic substances. Analyses for both plasma and urine were better by the present procedure. The mean and standard deviation for nickel in plasma from 15 healthy individuals was 2.13 +/- 0.58 microgram/liter by this method. For nickel in urine from 15 healthy men the mean and standard deviation was 4.45 +/- 1.9 microgram/liter. The coefficient of variation for plasma was 11.9%, and for urine 12.2% in 10 analyses of the same plasma and urine with the protein-precipitation procedure, as compared with 26.0 and 38.2%, respectively, by the acid-digestion technique.

Adult↗

[The properties of precipitation reactions between secretion products in the oviduct of pleurodeles: identification of a lectin].

The properties of two precipitation reactions occurring between secretory products from the oviduct of Pleurodeles waltl have been studied. It has been demonstrated that a lectin is involved in one of the reactions. This lectin precipitated glycogen and starch and required calcium; the most potent saccharide inhibitors were 2-amino-2-deoxy-D-glucose and D-glucose, respectively. The other reaction was related to glycoproteins (probably sulfated glycoproteins) that contained sulphur. The properties of this reaction were not the same as purely ionic interactions; basic protein-acidic polysaccharide interactions have been compared. A lectin was probably implicated but this could not be demonstrated because no saccharide inhibitor was found. There are several similitudes between this reaction and the lectin-galactoside reaction which occurs in the reaction between cortical granule content and egg jellies in anurans.

Animals↗