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Various birch-pollen and dog-epithelium allergen extracts were chromatographed using Sephadex G-75 and Sephadex G-200 columns. The chromatographic tractions were then tested by an in vitro paper disc radioallergosorbent test, using sera from allergen-sensitive patients. Calibrated columns were used to approximate molecular weights of some birch-pollen and dog-epithelium allergenic active components.

Birch pollen and dog epithelium allergen extracts were studied by ultracentrifugation in linear sucrose gradients. The various sucrose-gradient fractions were then tested for allergenic activity by the paper disc radioallergosorbent test, using sera from allergen sensitive patients.

Grass pollen allergens are shown to remain associated with protein material and a yellow pigment during paper chromatography and during dialyses and ultrafiltrations of various types. Dialysable* allergens comprise only a fraction of 1 per cent of the total activity and the amount of activity extractable by diethylene glycol (DEG) and similar solvents is of the same order.

A method has been developed for the assay of allergens and may be summarized as follows: 1. Filter paper discs are activated with BrCN. 2. The allergens are coupled to the reactive groups on the paper disc carrier. 3. IgE immunoglobulin, present in the sera of allergic patients, then attaches itself to the allergen coupled to the paper discs. 4. 125 I-labeled anti-IgE antibodies thereafter interact with the IgE molecules attached via the allergens to the paper carrier. Examples of the assay with the use of dog, cat, horse, cow, and timothy allergens are presented. The allergen assay method can conveniently be used to compare the "potency" of different allergen extracts and to check the extraction procedure, storage, and further treatment of allergens.

SUMMARY.— An investigation into the pathogenesis of the occupational skin disorder known as “tulip finger” showed it to be determined by an allergic mechanism. This is in line with the findings of some other investigators. Paper-chromatography of active extracts from tulip bulbs, and subsequent patch-tests with these paper-chromatograms in sensitized individuals, made it possible to localize the allergic principle in the paper-chromatograms and to find a reagent (Tollens) for chemical spot tests with the active agent. In later experiments we used the Tollens reaction to locate the active substance in paper-chromatograms, without doing patch tests. A quantity of “crude allergen” was obtained by preparative paper-chromatography. Qualitative analysis indicated a compound related to sugars, which is probably a glucoside. Hydrolysis of this compound yielded glucose, and a substance identified by chemical, spectroscopic and gas-chromatographic methods as: α-methylene-γ-butyrolactone.

An allergenically active basic protein, antigen Ra5, was isolated from DEAE-fraction C of the aqueous extract of short ragweed pollen by gel filtration and cation exchange chromatography. The antigen was homogeneous by a number of physicochemical and immunological criteria, and showed no cross-reaction with ragweed pollen antigens E, Ra3 and BPA-R. It was devoid of carbohydrate, had a molecular weight of 5,130 ± 50 in the analytical ultracentrifuge and the following composition: lys4 arg2 asp2 ser4 glu4 pro3 gly4 ala3 (cys/2)6 val3–4 ileu leu1–2 tyr3 try2.

The basic protein encephalitogen, without, prior denaturation, was rapidly digested with such proteolytic enzymes as trypsin, pronase, chymotrypsin, and pepsin; from 80–90% of the proteolysis occurred within 1 hr. The number of peptides released by these enzymes after 2 hr were about 28, 54, 20, and 14, respectively. Each proteolytic digest was chromatographed on Sephadex G-25 and G-75 in order to separate the peptides from large partially digested material. Only the peptide fractions obtained from the pepsin digestion possessed significant activity in inducing experimental allergic encephalomyelitis (EAE). One of the peptide fractions had higher encephalitogenic activity, on a weight basis, than either the original protein or the unfractionated digest. In contrast, the chymotryptic digest, while lacking encephalitogenic activity, possessed antibody-combining activity as shown by the Ouchterlony and passive hemagglutination-inhibition (PHI) test. The pepsin digest produced peptides which gave a negative Ouchterlony test. The tryptic peptides were negative in all tests—Ouchterlony, PHI, and EAE induction. These results suggest that the encephlitogenic and antibody-combining activities reside in separate or overlapping regions of the polypeptide chain. The retention of encephalitogenic activity in relatively small pepsin peptides further suggests that the EAE determinant may be defined by a region of the polypeptide chain lacking a folded tertiary structure.

Determination of 5 -hydroxy-indolacetic-acid ( 5 -HIAA) in urine and nasal mucus in provoked allergic rhinitis

Chlorogenic acid, a simple phenolic compound of low molecular weight, is a major allergenic constituent of green coffee dust, castor bean, and oranges. Patients with violent respiratory allergy to inhaled coffee dust and large positive intradermal reactions to chlorogenic acid are able to ingest coffee or other foods containing chlorogenic acid without difficulty. It was shown by means of paper chromatography and by ultraviolet absorption spectrophotometry that chlorogenic acid was transformed in vitro to a presumably nonallergenic derivative by secretions of the small intestine, whereas it remained intact when exposed to respiratory tract secretions. These findings offer a possible explanation in a particular case for the clinical observation that skin testing is a reliable diagnostic procedure for inhalant allergens but is unreliable for ingested allergens.

In 13 patients with allergic asthma disodium cromoglycate protected fully only two from an allergen-induced asthmatic attack. Inhalation of disodium cromoglycate did not improve lung function in five patients with long-standing chronic asthma. Previous clinical trials do not show convincing evidence that this drug improves bronchial asthma in a high percentage of cases, but it seems to help a small minority of patients.

This study was undertaken to examine the possibility that IgE is not the only immunoglobulin responsible for immediate allergic reactions. A group of asthmatics were investigated in whom immediate allergic reactivity of the bronchi to common inhalant allergens had been confirmed by provocation tests. Their sera were fractionated and the reaginic activity of the immunoglobulin classes was studied by passive cutaneous anaphylaxis testing in monkeys. The results showed that the immediate allergic reactions were due to IgE antibodies in most patients, but there was a group with reactions due to short-term anaphylactic IgG antibodies. It was not possible to inhibit the IgG-mediated responses with disodium cromoglycate. As these two groups had clearly different serum IgE levels the estimation of IgE provided an important guide to the management of these patients.

The NH 2 -terminal amino acid sequence of a major allergenic fragment (TM2) from cod was elucidated and shown to have the following structure: Tryptic hydrolysis of the molecule yielded 6–8 oligopeptides. No allergenicity was found for either of these peptides or for fragment TM 2 after the removal of the 17 NH 2 -terminal residues.

The amino acid sequence of the COOH-terminal fragment TM2 of CIK! allergen M has been determined by the dansyl-Edman procedure The fragment consists of 58 residues (mol.wt. 3,854). and the sequence shows great homology with the sequence of the corresponding fragment of white muscle albumin from carp. hake, and pike. Unique tor the TM2 (and allergen If) from cod is the presence of a tryptophyl residue at position 5 from the COOH-terminal end.

(1967). Dermatitis in the Forest Product Industries. Archives of Environmental Health: An International Journal: Vol. 15, No. 3, pp. 322-326.