Altered glycosylation, expression of serum haptoglobin and alpha-1-antitrypsin in chronic hepatitis C, hepatitis C induced liver cirrhosis and hepatocellular carcinoma patients.

Liver cirrhosis with hepatitis C viral infection (HCV-LC) causes high risk to develop hepatocellular carcinoma (HCC). Besides diagnosis of liver cirrhosis by biochemical test, imaging techniques, assessment of structural liver damage by biopsy shows several disadvantages. Our aim was to monitor the changes in the expression level of serum proteins and their glycosylation pattern among chronic hepatitis C (HCV-CH), HCV-LC and HCC patients with respect to controls. 2D gel electrophoresis of HCV-CH, HCV-LC and HCC patients' sera showed several protein spots, which were identified by LC-MS. The change in the expression of two prominent protein spots, haptoglobin (Hp) and alpha 1-antitrypsin (AAT) was evaluated by western blot and ELISA. The changes in glycosylation pattern of these serum proteins were assayed using different lectins. Increased level of Hp and AAT was observed in HCV-LC and HCC patients' group whereas those were found to be present less in HCV-CH patient groups with respect to control as determined by ELISA using monoclonal antibodies. Decreased level of sialylation in both Hp and AAT was observed in HCV-LC and HCV-CH patients' group whereas increased level of sialylation was observed in HCC patient groups by ELISA using Sambucus nigra agglutinin. On the other hand increased level of fucosylation in two serum glycoproteins was observed in HCV-LC and HCC patients' group using Lens culinarris agglutinin. High glycan branching was found in HCV-LC and HCC patient groups in Hp but not in HCV-CH as determined by Datura stramonium agglutinin. However, there was no such change observed in glycan branching in AAT of HCV-CH and HCV-LC patients' groups, to the contrary high glycan branching was observed in HCC patients' group. Increased level of exposed galactose in both serum proteins was observed in both HCC patients' group as determined by Ricinus communis agglutinin. The present glycoproteomics study could predict the progression of HCV-CH, HCV-LC and HCC without the need of liver biopsy.

Interplay of T Helper 17 Cells with CD4(+)CD25(high) FOXP3(+) Tregs in Regulation of Allergic Asthma in Pediatric Patients.

Background. There is evidence that Tregs are important to prevent allergic diseases like asthma but limited literature exists on role of TH17 cells in allergic diseases. Methods. Fifty children with asthma and respiratory allergy (study group) and twenty healthy children (control group) were recruited in this study. Total IgE levels and pulmonary function tests were assessed. The expression of Tregs and cytokines was determined by flow cytometry. Results. The average level of total IgE in study group (316.8 ± 189.8 IU/mL) was significantly higher than controls (50 ± 17.5 IU/mL, P < 0.0001). The frequency of TH17 cells and culture supernatant level of IL-17 in study group (12.09 ± 8.67 pg/mL) was significantly higher than control group (2.01 ± 1.27 pg/mL, P < 0.001). Alternatively, the frequency of FOXP3 level was significantly lower in study group [(49.00 ± 13.47)%] than in control group [(95.91 ± 2.63)%] and CD4(+)CD25(+)FOXP3(+) to CD4(+)CD25(+) ratio was also significantly decreased in study group [(6.33 ± 2.18)%] compared to control group [(38.61 ± 11.04)%]. The total serum IgE level is negatively correlated with FOXP3 level (r = -0.5273, P < 0.0001). The FOXP3 expression is negatively correlated with the IL-17 levels (r = -0.5631, P < 0.0001) and IL-4 levels (r = -0.2836, P = 0.0460). Conclusions. Imbalance in TH17/Tregs, elevated IL-17, and IL-4 response and downregulation of FOXP3 were associated with allergic asthma.

Activation dependent expression of MMPs in peripheral blood mononuclear cells involves protein kinase A.

Monocyte/Macrophages are integral cellular components of inflammation. Matrix metalloproteinases (MMPs) produced by these cells play a crucial role in every aspect of inflammation. Results of the investigations on activation dependent upregulation of MMPs in human peripheral blood mononuclear cells in culture using different lectins as an in vitro model system to mimic inflammatory monocytes are presented. Under normal physiological conditions the monocytes produced only very low amount of MMPs in an indomethacin insensitive PG/cAMP independent manner. Zymographic analysis and ELISA showed that treatment of monocyte with lectins like concanavalin A (ConA), wheat germ agglutinin (WGA) and Artocarpus lakoocha agglutinin (ALA) caused upregulation of MMPs and the maximum effect was produced by ALA. ALA significantly upregulated MMP-9 in a concentration and time dependent manner. Immunoblot analysis and RT-PCR confirmed ALA mediated upregulation of MMP-9 production. Inhibition of ALA effect by indomethacin and reversal of the indomethacin effect by Bt(2)cAMP indicated involvement of cAMP dependent signaling pathway. Further support for the prostaglandin mediated effect was obtained by the upregulation of cyclooxygenase by ALA. H-89, an inhibitor of protein kinase A (PKA), inhibited the expression of MMP-9 indicating that ALA mediated upregulation of MMP-9 is mediated through PKA pathway. Increase in MMP production and increase in cyclooxygenase activity and inhibition of the effect of ALA on MMP production by indomethacin suggested that the ALA activated monocytes in culture can be used as an in vitro model system to study the intracellular signaling process involved in the mediation of inflammatory response.

Ficus cunia agglutinin for recognition of bacteria.

Interaction of bacteria with lectin using anti-lectin antibody by ELISA is an established method. In the present study, we have devised a simple ELISA using a biotinylated lectin and antibiotin-HRP. Ficus cunia agglutinin (FCA), which has shown the specificity towards alpha/beta anomers of GlcNAc and other-NAc containing sugars like LacNAc and GlcNAcbeta(1-4/6)GlcNAc, was used as a model lectin for the study of interaction with immobilized microorganisms on ELISA plate. The bacterial cells of E. coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Bacillus subtilis and Staphylococcus aureus showed binding with FCA and the degree of binding was dependent on the bacterial surface antigen. This method is considered a simple technique to study the lectin-bacteria interaction.

Changes in the allergenicity during different preparations of Pomfret, Hilsa, Bhetki and mackerel fish as illustrated by enzyme-linked immunosorbent assay and immunoblotting.

BACKGROUND: Although the identification and characterization of several fish allergens have already been reported, there is almost no data on Indian fish allergens and the effect of thermal processing on their allergenicity. This study aimed at the evaluation of the changes in the level of allergenicity of 4 highly consumed Indian fishes, i.e. pomfret, hilsa, bhetki and mackerel, that occurred after boiling and frying. METHODS: In this study 110 patients with fish hypersensitivity as evidenced by clinical history and symptoms were recruited based on their positive skin prick test results. The raw, boiled and fried muscle extracts of the 4 fishes were prepared, and each extract was tested by ELISA and immunoblotting with patients' sera. RESULTS: ELISA and immunoblotting studies demonstrated that the raw muscle extracts of pomfret, hilsa, bhetki and mackerel were allergenic. While the allergenicity of boiled and fried extracts of pomfret and hilsa was considerably reduced, maximum allergenicity of bhetki was demonstrated in the fried extract. The degree of allergenicity of bhetki was demonstrated in the order fried>boiled>raw while that of mackerel followed the order raw>boiled approximately fried. CONCLUSION: The specific IgE-binding activity and immunoblot profile clearly showed that pomfret and hilsa fish allergens are heat-labile, while allergens of bhetki and mackerel maintained strong reactivity even after thermal treatment.

Mucin binding mitogenic lectin from freshwater Indian gastropod Belamyia bengalensis: purification and molecular characterization.

A lectin was purified from the hemolymph of the freshwater Indian gastropod Belamyia bengalensis. The purification involved successive ion-exchange chromatography on Resource Q and gel filtration on Superose 12 column in FPLC system. Homogeneity of the protein was confirmed by polyacrylamide gel electrophoresis. Belamyia bengalensis lectin (BBL) was a monomeric protein with a molecular weight of 33 kDa as demonstrated by gel filtration and SDS-PAGE. It is a glycoprotein containing 6% total sugar and its activity is highly dependent on Ca(2+). BBL agglutinated human erythrocytes and is a blood group non-specific lectin. It agglutinated animal erythrocytes also. Hapten inhibition studies indicated that BBL shows binding specificity only for N-acetyl-D-glucosamine and N-acetyl-D-galactosamine at a high concentration among the mono- and oligosaccharides tested. Among the glycoproteins used for hemagglutination-inhibition assay, porcine submaxillary mucin was found to be the best inhibitor. Chemical modification studies indicated that Lys, Arg, and Trp are essential for the sugar-binding activity of BBL. Circular dichroism spectra revealed high content of alpha-helical structure in the lectin. BBL is a potent mitogen as it stimulated the T-lymphocyte proliferation, specifically the Th1 subset.

Purification and characterization of an extracellular agglutinin from Tricophyton rubrum with specificity towards sialic acid containing glycoconjugates.

An agglutinin, a monomeric glycoprotein with a molecular mass of about 6.5 kDa and containing 18% sugar has been purified to an apparent homogeneity from a 21 days old culture filtrate of an anthropophilic dermatophyte Tricophyton rubrum. It is a human blood group non-specific agglutinin which also agglutinates animal erythrocytes and Ehrlich ascites carcinoma and Sarcoma-180 cells. It is thermally stable and exhibits maximum activity at pH 8. Amino acid analysis shows a significant amount of glycine, with no cysteine. Glycoproteins inhibited the hemagglutination of the agglutinin, but not the simple sugars, including sialic acid. Fetuin is the most potent inhibitor among the glycoproteins tested. This inhibition gives a hint to binding with Galbeta1-3GalNAc or Galbeta1-4GlcNAc residue containing sialic acid at the terminal position with alpha 2-6 or alpha 2-3 linkage.

Serodiagnosis of ascariasis with specific IgG4 antibody and its use in an epidemiological study.

In an earlier study Ascaris-specific IgG4 antibody was found to be elevated in cases of ascariasis. However, the usefulness of the elevated levels of this antibody in Ascaris infection as a diagnostic marker has not been well established. In India, in early 1999, blood samples of 83 cases of Ascaris infection, 35 cases of other nematode infection and 53 control subjects (without any helminth infection) were tested for anti-Ascaris IgG4 by ELISA. Further anti-Ascaris IgG4 levels in the blood of Ascaris-infected patients were determined, after eradication of the worms with drugs, at regular intervals to ascertain the duration of elevation of titre of the serological marker following initial infection. This information would indicate the sensitivity of the test as a diagnostic marker for recent infection. Blood samples of 422 rural people were also tested for anti-Ascaris IgG4 titre to ascertain the prevalence of ascariasis in the community. High levels of anti-Ascaris IgG4 antibody (OD 1.246 +/- 0.212) were found in all the 83 Ascaris-infected subjects compared to controls (OD 0.158 +/- 0.047). Anti-Ascaris IgG4 antibody levels of other nematode-infected subjects were comparable to the controls. Anthelmintic treatment of 8 Ascaris-infected subjects caused sequential fall of IgG4 level in their blood, and its titre reached control level within 6 months of deworming. Of 422 individuals from the rural community 229 (54.3%) had significantly high levels of specific IgG4 antibody against Ascaris excretory-secretory antigen, suggesting that they were infested with Ascaris. Thus, this study demonstrated that anti-Ascaris IgG4 antibody is a very sensitive and specific marker for the diagnosis of Ascaris infection. Utilizing this test, a significant number of a rural population could be diagnosed with Ascaris infection in West Bengal, India.

Possible approach for serodiagnosis of ascariasis by evaluation of immunoglobulin G4 response using Ascaris lumbricoides somatic antigen.

Somatic antigen of Ascaris lumbricoides was purified to homogeneity (molecular mass, 34 kDa) by ammonium sulfate fractionation and successive chromatographic procedures, namely, gel permeation, ion exchange, and high-performance gel permeation liquid chromatographies. The antigen showed strong binding with immunoglobulin G (IgG) in Ascaris-infested patients and was cross-reactive with IgE and IgG in patients infested with other nematodes. It reacted specifically with IgG4 (P < 0.001) in 63 Ascaris-infested patients, which represented 65% of the total IgG response, though cross-reactivity with IgG1, IgG2, and IgG3 subclasses was observed, indicating the unique specificity of this test system and its potential utility in the serodiagnosis of ascariasis.

Heart rate variability in dilated cardiomyopathy.

Chronic heart failure is associated with excessive neurohormonal activation. Analysis of heart rate variability is considered a valid technique for assessment of the autonomic balance of the heart. Twenty symptomatic patients of dilated cardiomyopathy in NYHA class II-IV symptomatic status and as many normal controls were subjected to 24 hours Holter monitoring to assess the heart rate variability with both time domain and frequency domain analysis. Age of the patients ranged from 12 to 67 years (mean +/- SD 38.6 +/- 7 years), the male-female ratio was 4:1. The left ventricular ejection fraction of the patients was between 18-42 percent (mean +/- SD 30.2 +/- 9%) and all received diuretics, digoxin and angiotensin-converting enzyme inhibitors. Heart rate variability parameters measured included mean heart rate with standard deviation, hourly heart rate with SD and the mean of all normal RR intervals from the 24-hour recording. Time domain measures calculated were SD of all normal RR intervals, SD of 5 minute mean RR intervals and root mean square of difference of successive RR intervals. Using spectral plots, frequency domain subsets of low frequency and high frequency were analysed and expressed in normalised units. Total power was also measured. In the dilated cardiomyopathy patients, mean 24-hour heart rate in beats per minute was significantly higher in comparison to controls (82 +/- 13 vs 72 +/- 8; p < 0.001) whereas mean hourly heart rate with standard deviation (msec) was significantly lower (97 +/- 41 vs 232 +/- 25; p < 0.001), SD of all normal RR intervals (msec) was 85.5 +/- 26.3 vs 139.4 +/- 16.9 in controls (p < 0.001), SD of 5 minute mean RR intervals (msec) was also significantly less in patients in comparison to controls (75.8 +/- 39.6 vs 130.8 +/- 20.3; p < 0.001). However, although root mean square of difference of successive RR intervals (msec) was reduced in patients (30.1 +/- 9.3 vs 37.3 +/- 11.7; p < 0.05), the difference was non-significant. Low frequency power (0.05-0.15 Hz) (normalised units) was reduced in the dilated cardiomyopathy group (0.0721 +/- 0.003 vs 0.136 +/- 0.047 in the control group; p < 0.001). High frequency power (0.35-0.50 Hz) (normalised units) (0.08 +/- 0.05 in patients vs 0.09 +/- 0.02 in controls; p > 0.1) and total power frequency (0.02-0.50 Hz) (normalised units) (0.34 +/- 0.05 in patients vs 0.35 +/- 0.12 in controls; p > 0.1) was non-significantly different in the two groups. Regression analysis showed a significant decrease in SD of all normal RR intervals, SD of 5 minute mean RR intervals, low frequency, high frequency, total power and a non-significant decrease in root mean square of difference of successive RR intervals with a decrease in ejection fraction percent whereas there was a significant decrease in SD of all normal RR intervals, SD of 5 minute mean RR intervals, low frequency and total power and a less significant decrease in root mean square of difference of successive RR intervals and high frequency power with an increase in NYHA class. At 6 months duration, 6 patients were lost to follow-up, 3 patients were readmitted (2 for congestive cardiac failure, one of paroxysmal supraventricular tachycardia). One patient who was NYHA class IV at baseline was readmitted for congestive cardiac failure and showed much lower heart rate variability parameters compared to the average of the patients. We conclude that in symptomatic dilated cardiomyopathy patients, heart rate variability parameters are significantly reduced in comparison to control subjects.

Determination of the immunodominant part in the O-antigenic polysaccharide from Escherichia coli O128 by ELISA-inhibition study.

The immunodominant part in the O-antigenic polysaccharide from Escherichia coli O128 was immunologically characterized by an enzyme-linked immunosorbent assay (ELISA). The antibody specificity was determined by the inhibitory effects of the methyl glycosides of constituent mono- and oligosaccharides synthesized related to the O-antigenic polysaccharide from E. coli O128. It was found that methyl alpha-L-fucopyranoside was the most effective inhibitor amongst the monosaccharides while the highest antibody specificity was directed towards the trisaccharide with the structure: beta-D-GalpNAc-(1-->6)-[alpha-L-Fucp-(1-->2)]-beta-D-Galp-1-->OMe suggesting that the monospecific antibody has the extended combining site.

Saracin: A lectin from Saraca indica seed integument induces apoptosis in human T-lymphocytes.

Saracin, a seed integument lectin from Saraca indica is highly specific for binding N-acetyl-neuraminyl-N-acetyllactosamine [Neu5Ac-alpha-(2-6)/(2-3)-D-Gal-beta-(1-4)-D-GlcNAc]. This lectin has been found to be mitogenic for human lymphocytes, and this mitogenic activity could be inhibited in presence of fetuin. Further, treatment with saracin could induce secretion of IL-2 in a culture of resting human peripheral blood mononuclear cells (PBMC) after 48 h. Saracin has a higher affinity for the CD8(+) than CD4(+) T cells as revealed by FACS analysis. Agarose gel electrophoresis of DNA isolated from lymphocytes cultured under different conditions has shown that this lectin could induce apoptosis in activated T-lymphocytes, as also confirmed by flow cytometric studies. Phenotypic analysis of the apoptotic cells reveals that they belong to CD8(+) T cells lineage. Four surface glycoproteins of PBMC have been found to interact with saracin in a trisaccharide [Neu5Ac-alpha-(2-6)/(2-3)-D-Gal-beta-(1-4)-D-GlcNAc]-sequence specific manner. Saracin seems to be an interesting immunomodulator for the mammalian immune system.

Inhibition of bacterial respiration by a low-molecular weight lectin, scyllin, from Scylla serrata crab hemolymph.

Interaction of plant and/or invertebrate lectins with mammalian cells and different microorganisms is well known. In the present study, we have demonstrated that scyllin, a low molecular weight (MW 4000) lectin from the edible crab Scylla serrata hemolymph, purified by GalNAc-Sepharon affinity column followed by Mono-Q ion exchanger in FPLC exhibits antimicrobial activity against Bacillus cereus and Escherichia coli by inhibiting endogenous respiration as well as exogenous glucose oxidation. In both the cases oxygen consumption has been measured in an oxygraph. Scyllin has produced 50% inhibition of endogenous respiration at a concentration of 110 micrograms/ml and 125 micrograms/ml in B. cereus and E. coli respectively. It also reduced the exogenous glucose oxidation by 50% at a concentration of 12 micrograms/ml and 80 micrograms/ml respectively in B. cereus and E. coli. From the above study the mechanism of bacterial growth inhibitory property of scyllin is suggested though the other studies such as inhibition of nucleic acid biosynthesis, cell wall biosynthesis etc. to evaluate its total mode of inhibitory action are not yet obtained.

Evaluation of IgG4 response in ascariasis by ELISA for serodiagnosis.

The excretory/secretory (ES) antigen(s) of Ascaris lumbricoides was fractionated into 10 fractions by gel chromatography on a Suparose 12 column in FPLC. Of these, the third fraction (Al III), showing binding activity with both IgE and IgG antibodies of A. lumbricoides infected patients' sera, was further resolved into 2 fractions (Al IIIa and Al IIIb) on passage through a Mono Q column. Al IIIb was found to be the most potent antigen due to its high binding affinity with IgE and IgG antibodies of Ascaris infected patients as evidenced by ELISA inhibition. Although a two to five-fold increase of serum IgE level was observed in all helminthic parasite infected patients studied compared to control subjects, Al IIIb specific IgE was detected in sera of all Ascaris infected and only 40% of hookworm infected patients. When Al IIIb was tested by ELISA with sera of control subjects, Ascaris, hookworm, Strongyloides and Trichuris infected patients, strong binding was observed with the IgE and also IgG of all the Ascaris infected patients; however, it cross-reacted with IgG in 50% of hookworm, 28.6% of Trichuris trichura and 22.2% of Strongyloides infected patients' sera; but with IgE only in 40% of hookworm infected patients' sera. Further study showed specific detection of IgG4 in all the serum samples of 65 Ascaris infected patients when Al IIIb antigen was allowed to react with different subclasses of IgG by ELISA, giving a sensitivity of 100%. Reactivities of Al IIIb with IgG1 and IgG3 were only 47.6 and 11.8% respectively and there was no reactivity with IgG2 subclass. No IgG4 reactivity against Al IIIb was observed in the sera of hookworm, Trichuris or Strongyloides infected patients and was similar to that observed with control subjects showing the 100% specificity of the test system. This study may therefore be regarded as a novel technique for serodiagnosis of ascariasis by measuring Ascaris specific IgG4.

Subcellular distribution of jacalin in Artocarpus integrifolia seed.

Seeds of Artocarpus integrifolia (jack fruit) contain large amounts of the anti-T lectin, jacalin. The mature seeds of jack fruit were homogenized in 0.25 M sucrose and separated by differential centrifugation into four fractions, viz wall, intermediate, and microsomal pellets and soluble supernatant. The lectin activity was associated with the wall pellet collected at low speed centrifugation. The other three fractions obtained by centrifugation at gradually higher speeds contained a similar lectin but of very low specific activity. The distribution pattern of jacalin remained unchanged in the presence of EDTA and/or Triton X-100 indicating that the lectin was not membrane bound. Immunofluorescent staining of jack fruit seeds showed that jacalin was localized in the cell wall in the intracellular space, which corroborated the results of fractionation studies. The possible relevance of these results to the function of lectin in the plant cell is discussed.

Saracin: a lectin from Saraca indica seed integument recognizes complex carbohydrates.

A lectin isolated from Saraca indica seed integument was purified by affinity chromatography on porcine thyroglobulin Sepharose followed by Sephadex G-50 and shown to be homogeneous by PAGE. It showed a single band on SDS-PAGE in the absence and presence of 2-mercaptoethanol corresponding to a M(r) of congruent to 12,000, thus indicating it to be a monomer. The lectin agglutinated erythrocytes of human A, B, O and AB blood group, animal erythrocytes as well as Ehrlich ascites cells. It is a thermostable glycoprotein containing 11.6% carbohydrates and large proportions of acidic amino acids. In haemagglutination-inhibition assays, among the tested glycoproteins, porcine thyroglobulin having the NeuAc alpha (2-6)/(2-3)D-Gal beta (1-4)D-GlcNAc sequence was found to be the most potent; however, its asialo counterpart was non-inhibitory. The lectin is present solely in the seed integument even in the immature stage. During maturation of the seed the lectin activity declined and was completely absent when totally matured and dried. Studies in vitro showed that on incubation at 37 degrees the seed gradually lost its lectin activity which was completely absent after 62 days with 88.5% loss of water. Similar studies on scraped seed integument revealed that the lectin activity was lost in 22 days with 87.5% dehydration.

Cocos nucifera pollen inducing allergy: sensitivity test and immunological study.

Among the heterogeneous population (n = 975) in greater Calutta, sensitization to Cocos nucifera pollen accounts to be 2.65% and for atopic patients (n = 204) 47.06%. Out of 24 patients who had C. nucifera pollen sensitivity and suffered from asthma and allergic rhinitis, 16 showed sensitivity also to other allergens. All were skin test positive and 19 of them were phadezym RAST positive to C. nucifera pollen extract. Bronchial provocation test appeared to be positive in 7 out of 8 patients included in the test and no late response or non-specific reactions were observed. C. nucifera pollen extract on fractionation by ion-exchange chromatography following gel filtration yielded two major allergenic protein fractions, CnII (M(r) 158,000) and CnVII (M(r) 2900) as evidenced by skin prick test, ELISA-inhibition and immunoblot analysis. Hence, C. nucifera pollen should be considered to be a relevant allergen and thus included in the panel of allergens for routine clinical use.

Typing of Shigella dysenteriae strains of different serogroups by lectins.

Ten different serogroups of Shigella dysenteriae were typed with the aid of lectins of known sugar specificity resulting from their interactions with the carbohydrates on lipopolysaccharides in the outer membrane of bacteria as evidenced by the agglutination-inhibition assay with simple carbohydrates. Lipopolysaccharides of two serogroups of Shigella were precipitated with different lectins and the results were corroborated by those derived from the agglutination assay suggesting that Shigella dysenteriae can be characterized on the species level with the aid of lectins.

Isolation and characterization of two IgE-reactive proteins from Azadirachta indica pollen.

Two allergenically active components present in the Azadirachta indica whole pollen extract have been isolated by sequential ammonium sulfate precipitation (0-90%), DEAE-Sephadex A-50 ion-exchange chromatography followed by gel filtration through Sephadex G-200. The allergenicity of fractionated materials has been tested by skin prick test and ELISA inhibition which reveal that AIaI and AIaIVb are the major allergens. Immunoblot confirms the IgE-binding activity of the proteins. Although both fractions are found to be homogeneous by SDS-PAGE, isoelectric focusing produces more than one isoelectric point in AIaI (pI = 3.15, 3.3 and 3.5) and AIaIVb (pI = 6.0 and 6.2). Amino acid analyses of the two allergens, the effect of pH on them and cross-reactivity between them have been discussed.

Placebo-controlled immunotherapy with Cocos nucifera pollen extract.

The effectiveness of Cocos nucifera pollen extract immunotherapy (CPE-IT) was studied in 96 patients allergic to C. nucifera pollen. A placebo-controlled study was performed at random for a period of 6-12 months. The clinical status of the patients measured by the symptom-medication scores demonstrated that C. nucifera pollen-allergic patients had significant (p < 0.005) clinical improvement after CPE-IT in comparison to placebo treatment. Serological study resulted a significant reduction (p < 0.001) of specific IgE and significant elevation (p < 0.01) of specific IgG in post-therapeutic patients' sera which were correlated significantly (r = 0.45, p < 0.001); the changes of the above immunoglobulin levels in the placebo-treated patients were nonsignificant. However, there was no correlation between symptom-medication scores and changes in specific serum IgE or IgG levels.