Timothy Grass Pollen Extract Research Articles

381 Application of the first international reference extract of timothy grass ( [formula omitted]) pollen for the standardisation of commercial allergen extracts

381 Application of the first international reference extract of timothy grass ( [formula omitted]) pollen for the standardisation of commercial allergen extracts

The international collaborative study establishing the first international standard for timothy ( Phleum pratense) grass pollen allergenic extract

A selected candidate international reference preparation of timothy grass (Phleum pratense)-pollen extract was studied together with two other freeze-dried timothy pollen allergenic extracts in a multinational study. The collaborators used RAST inhibition, histamine release, quantitative immunoelectrophoresis (crossed immunoelectrophoresis/crossed radioimmunoelectrophoresis and rockets), isoelectric focusing, and other methods. The total allergenic potencies measured in RAST inhibition were evaluated for validity of linearity and parallel-line response. The relative concentrations of some important individual allergenic components were measured. The relative potencies for the total allergenic activity and the timothy components studied in each preparation were expressed relative to the selected candidate. This preparation was established in 1983 by the World Health Organization expert committee on biologic standardization as the international standard for timothy grass pollen extracts with assigned units of 100,000 IU per ampule.

IgG subclass antibody response in grass pollen-allergic patients undergoing specific immunotherapy. Prognostic value of serum IgG subclass antibody levels early in immunotherapy.

All four subclasses of IgG antibodies to timothy grass pollen extract were measured by a three-layer immunoradiometric assay in sera from 20 grass pollen-allergic patients who underwent specific immunotherapy in a 3-year prospective study. Both IgG1 and IgG4 antibody levels rose significantly during the first 8 weeks of immunotherapy. IgG1 antibody level passed its peak (median 5.4 U/ml) after 12 weeks. At this time, the ratio between the medians of IgG1 and IgG4 antibodies was 2.25. IgG4 antibody level reached its peak (median 11.6 U/ml) just before termination of immunotherapy. At this time IgG1/IgG4 ratio was 0.43. Two years after the end of immunotherapy, IgG1 and IgG4 antibody levels were 0.0 and 1.8 U/ml in median, respectively. The amounts of IgG2 and IgG3 antibodies detected in the sera were less than 1.6 U/ml and were considered insignificant. Preseasonal serum IgG1 and IgG4 antibody levels did not correlate significantly with symptom scores in the subsequent season. Serum IgG4 level obtained after 12 weeks of immunotherapy was significantly correlated to symptom score in the third season, i.e. the season just after termination of therapy (rs = 0.529, t = 2.567, P = 0.02). In this work, a serum IgG4 antibody level higher than 8.0 U/ml after 12 weeks of therapy predicted poor clinical result at the end of immunotherapy with 100% sensitivity and 87% specificity. An IgG4/IgG1 ratio greater than 1.0 after 12 weeks' therapy had the same predictive value.

An automated particle counting immunoassay (PACIA) for determination of blocking antibodies against timothy grass pollen in sera from desensitized allergics.

An automated particle counting immunoassay (PACIA) for measurement of blocking antibodies (antigen neutralizing capacity) against timothy grass pollen extract in sera from desensitized allergics is described. Latex particles coated with F(ab')2-anti-timothy are agglutinated by timothy. Serum containing anti-timothy antibodies inhibits the agglutination. Non-agglutinated particles are counted in a modified AutoCounter. Nineteen of 20 sera from timothy allergics who had undergone immunotherapy with purified timothy extract for 30 weeks, showed significant agglutination-inhibition. None of 42 normal human sera gave significant inhibition. The inhibiting antibody could be removed by absorption with protein A and was thus of non-IgE nature, i.e. blocking antibody. The results obtained correlated statistically significantly with those found with a double-antibody method (rS = 0.62, n = 20, t = 3.35, P less than 0.01) and with the cumulated dosage of timothy allergen extract administered to the individual patient (rS = 0.56, n = 20, t = 2.87, P less than 0.02). Between-assay coefficient of variation was from 6.4% to 18.3%. The capacity is 40 samples per hour. The method has also been applied to measurement of blocking antibodies to honey bee and wasp venom.

Antigenic and allergenic changes during storage of a pollen extract

The changes that occur in the antigens and allergens of a timothy grass pollen extract during storage were measured by radioallergosorbent test (RAST) inhibition, isoelectric focusing, (IEF), crossed immunoelectrophoresis (CIE) and crossed radioimmunoelectrophoresis (CRIE) and a dye-binding protein assay. Storage intervals varied from 7 to 92 days at temperatures of 4 °, 22 °, and 35 °C and the freeze-dried extract was rehydrated and stored in either phosphate-buffered saline with 50% glycerol (PBSG) or Coca's solution (CS). For extracts stored in CS at 35 °C, allergenic activity as measured by RAST inhibition dropped by almost one half at 7 days and certain of the IEF, CIE, and CRIE bands were no longer visible, showing that these properties correlated with allergenic activity. After 65 days, no IEF bands were visible but three CIE bands could be visualized. At 22 ° the changes were slower and at 4 °C no changes were observed for the 92-day storage time. Extracts stored in PBSG showed some of the same changes but were much more stable at all temperatures. During storage, the protein content as measured by dye binding was also reduced, showing that proteins were fragmenting. Because some of the allergenic proteins disappeared more rapidly than others, there was not only a loss in total allergenic activity but also a change in specificity. This raises serious questions concerning the use of such extracts, especially for diagnosis.

Clinical correlations between long-term (IgE) and short-term (IgG S-TS) anaphylactic antibodies in atopic and 'non-atopic' subjects with respiratory allergic disease.

The sera of atopic and non-atopic persons with allergic pulmonary disorders were examined for long-term sensitizing, IgE and short-term sensitizing heat-stable (S-TS) antibodies which were present separately or together in the sera of some patients sensitive to antigens such as budgerigar serum proteins and Aspergillus funigatus. In fourteen atopic patients with extrinsic asthma, six had both types of antibody to common allergens, and of nine non-atopic patients with crytogenic (intrinsic) asthma, four had only heat-stable short-term sensitizing antibodies. The sera of atopic subjects with type I prick test reactions and positive RAST's, showed specific IgE antibody by baboon PCA tests to budgerigar serum proteins, A. funigatus, Timothy grass pollen extract and hen egg extract, and not to Dermatophagoides farinae, possibly because of naturally occurring mite antibodies in the baboon. The sera of non-atopic asthmatics, who had given negative prick test but positive immediate, dual or late intracutaneous tests, and only late asthmatic reactions, contained precipitins in most cases and gave little or no RAST reaction. On baboon PCA these sera contained either, S-TS antibody alone, or S-TS plus long-term sensitizing antibody, or long-term sensitizing antibody alone. Some of the sera with long-term sensitizing antibody contained blocking antibody which could diffuse away in the 24 hr delay for the baboon PCA test and could also be responsible for the negative RAST. Tests with insoluble anti-IgE immuno-adsorbents on two sera from persons sensitive to aspergillus confirmed that the S-TS activity was not due to IgE, and on two sera with negative RAST and negative prick tests to budgerigar serum antigens confirmed that the 24 hr monkey PCA responses were due to IgE.

Immunological studies in vitro and in vivo of children with pollenosis given immunotherapy with an aqueous and a glutaraldehyde-treated tyrosine-absorbed grass pollen extract.

Ten patients treated with an aqueous Timothy grass pollen extracts and given monthly maintenance doses were compared to ten patients given a glutaraldehyde-treated tyrosine-adsorbed grass pollen extract as five weekly injections pre-seasonally. There were four non-treated controls. Both treated groups showed equivalent increases of total IgE, Timothy-specific IgE, and IgG, but the concentrations of Timothy-specific IgG antibodies were sustained only in the patients receiving maintenance therapy. An increase of IgE and RAST titres during the pollen season were obtained in all three groups. No consistent changes were seen in the results of leucocyte histamine release tests and naso-conjunctival provocation tests from spring to autumn. A Sepharo-protein A technique was used for the estimation of Timothy-specific IgG antibodies. The results obtained with this technique correlated excellently with those of the Farr technique and a double-antibody technique.

Chemical modification of crude timothy grass pollen extract. II. Class and specificity of antibodies induced by chemically modified timothy grass pollen extract.

The effects have been studied of three different chemical modifications of timothy grass pollen extract on various immunological properties. The ability to induce IgG antibody with specificity for native antigen was least affected by glutaraldehyde treatment; IgE antibody production was reduced to a similar extent by all three modifications; there was no increase in IgM production; delayed reactions were reduced. New antigenic determinants were introduced by all the modifications, but the effect was minimal following glutaraldehyde treatment. The significance of these results and the potential application of modified allergen in hyposensitisation therapy are discussed.

Acetylsalicylic acid intolerance.

Abstract. Twenty‐seven ASA‐intolerant individuals were investigated. Twenty‐six of them suffered from chronic bronchial asthma, one from urticaria. All reacted to the ingestion of a therapeutic dose of ASA with more or less violent signs and symptoms resembling a systemic allergic (anaphylactic) reaction of the immediate type. Seventy percent of them reacted in the same way to one or more chemically unrelated drugs. Seventeen of these patients were skin‐tested with N‐salicyloyl‐ and N‐acetylsalicyloyl‐PLL without the appearance of immediate type reactions. Furthermore, the sera of these patients were passively transferred to rhesus monkeys. The intradermally sensitized monkeys were challenged by i.v. or intradermal injection of N‐salicyloyl‐BSA or N‐acetylsalicyloyl‐PLL or by oral administration of ASAN or ASA. No positive reaction was obtained. Intracutaneous sensitization of rhesus monkeys with the sera of individuals allergic to Timothy grass pollen revealed strong reactions of the immediate type following challenge with Timothy grass pollen extract. The IgE levels and the concentrations of the other immunoglobulins of the ASA‐intolerant individuals were found to be within the normal range. Thus, homocytotropic antibodies with N‐salicyloyl or N‐acetylsalicyloyl specificity could not be detected in the ASA‐intolerant individuals. These results, the frequency of intolerance to other drugs, the familial occurrence of ASA intolerance and the demonstration of “latent” ASA intolerance in about 20% of chronic asthmatics indicate that the clinical picture of the syndrome is not a sufficient criterion of its allergic origin. In our opinion the syndrome is caused by direct (i.e. not antibody mediated) liberation of histamine and other mediators by the offending drugs, due to an “inborn error of metabolism” in intolerant individuals. The syndrome should be properly denoted as ‘drug‐induced anaphylactoid syndrome“.

Immunological Properties of Carbohydrate and Protein Fractions of Timothy Grass Pollens

The effect of hot aqueous 45% phenol on timothy grass pollen extracts has been studied. Following separation into aqueous and phenolic phases, allergenic activity was recovered in the protein material in the phenol phase after precipitation with alcohol. The largely carbohydrate fraction recovered from the aqueous phase was non-allergenic. The methods used to test for allergenicity were prick-testing in the skin of allergic subjects and measurement of release of histamine from sensitised human leucocytes following challenge <i>in vitro. </i>In rats injected with <i>Bordetella pertussis, </i>homocy to tropic antibody was induced by injection of the protein fraction, but the carbohydrate fraction showed no such response.

Immunological properties of carbohydrate and protein fractions of Timothy grass pollens.

The effect of hot aqueous 45% phenol on timothy grass pollen extracts has been studied. Following separation into aqueous and phenolic phases, allergenic activity was recovered in the protein material in the phenol phase after precipitation with alcohol. The largely carbohydrate fraction recovered from the aqueous phase was non-allergenic. The methods used to test for allergenicity were prick-testing in the skin of allergic subjects and measurement of release of histamine from sensitised human leucocytes following challenge in vitro. In rats injected with Bordetella pertussis, homocy to tropic antibody was induced by injection of the protein fraction, but the carbohydrate fraction showed no such response.

Reverse passive cutaneous anaphylaxis in guinea pigs using extract of timothy grass pollen

Reverse passive cutaneous anaphylaxis in guinea pigs using extract of timothy grass pollen

Preliminary chemical investigations of a haptenic fraction isolated from Timothy grass pollen extracts

Preliminary chemical investigations of a haptenic fraction isolated from Timothy grass pollen extracts

Preliminary chemical investigations of a haptenic fraction isolated from Timothy grass pollen extracts

Preliminary chemical investigations of a haptenic fraction isolated from Timothy grass pollen extracts

Isolation and Immunochemical Properties of Haptenic Material from Timothy Pollen

Summary The isolation of a fraction (WST-Fr2) from the dialysate of timothy grass pollen extract is described. This fraction (WST-Fr2) appears homogenous by chromatography on Sephadex-G25, Amberlite IRC-50 and electrophoresis, but immunologically heterogenous as demonstrated by inhibition of several lines of precipitation in agar diffusion. WST-Fr2 possesses haptenic properties as demonstrated by its inhibition of specific precipitation and inhibition of the formation of lines of precipitation in agar diffusions. The results indicate the importance of considering the possible presence and significance of haptenic material in pollen extracts for laboratory studies as well as for clinical use.