CORRESPON DENCE
that for the purpose of performing the reproducibility experiments the serum To the editor: I have several comments was stored at 40C for several weeks. concerning the conclusions reached by I would not advocate such storage conW.B. Chodirker and A. Piotrowska in ditions for this type of experiment since their article entitled "Allergy testing: these, in my experience, have resulted comparison of skin and in vitro tests in similar inconsistent responses. For of allergic reagin" (Can Med Assoc J repeated testing we store serum in small 116: 1254, 1977) as one who has been aliquots at -200C. Chodirker and Piotrowska state that involved with several of the procedures mentioned in the article, particularly the study by Renoux and colleagues3 showed that rat mast cells did not bind the rat mast cell test (RMCT). The authors state that "the RMCT human IgE, but in fact the study was found to be poorly reproducible showed the opposite. Furthermore, and unreliable." While I do not defend Renoux and colleagues used three inthe clinical usefulness of the RMCT dependent techniques to demonstrate as a commercial test to detect imme- binding of* human IgE to rat mast cells. diate hypersensitivity reactions, I be- In addition, Plaut, Lichtenstein and lieve that the conclusions drawn by the Bloch4 actually showed that there was authors do not reflect the scientific a lack of histamine release from rat validity of the test. I have shown pre- mast cells incubated with ragweed alviously that there is a good correlation lergic serum and challenged with ragbetween the RMCT, the radioallerg&- weed antigen, and that this had nothing sorbent test (RAST) and intradermal to do with morphologic changes in rat skin tests.1 A double-blind study was mast cells. Moreover, several investigadone with serum from birch-allergic tors not mentioned in Chodirker and patients. Dr. S.G.O. Johansson of Piotrowska's article have demonstrated Sweden performed RAST analysis and histamine release from rat mast cells..7 Chodirker and Piotrowska imply that sent Dr. K. Ishizaka of Baltimore, Maryland nine samples of serum from I was involved in their study on a both atopic and nonatopic individuals. continuing basis. Although I was inDr. Ishizaka coded the samples and volved initially I cannot be held resent them to me for RMCT determina- sponsible for their experimental contions. An excellent correlation was ditions and conclusions. Furthermore, I believe that articles of this nature found between these in vitro tests. discourage further development of the Table III of the article in the Journal shows that only one of seven serum RMCT, which seems promising for the samples from atopic, highly sensitive, detection of IgE specific for a variety RAST-positive patients gave responses of drugs. It is well recognized that bindin the RMCT, whereas for three of five ing of human IgE by rat mast cells patients with moderate sensitivity the is weaker than binding in the homocyresponses in the RMCT were positive. totropic system; experimental condiThese results are similar to those found tions have to be exact for proper in another of my studies, which showed response in the test. Further work on that a significantly lower allergen con- mast cells from various strains of rats centration is required for serum with a or mice, and investigation into approhigh concentration of IgE antibody and priate storage conditions for these cells, that more allergen is needed for re- may ultimately resolve some of the sponse with lower concentrations of problems encountered with the RMCT. L. PERELMUTTER, PH D antibody.1 It is possible that the reacChief, hematology tions reported in Table III for the highBureau of clinical chemistry and hematology Health protection branch ly sensitive patients were performed in Health and Welfare Canada Ottawa, Ont. antigen excess, when such responses would be inhibited. To the editor: Dr. Perelmutter indicates As to the reproducibility of the that our conclusions do not reflect the RMCT, Chodirker and Piotrowska state scientific validity of the RMCT. We
Allergy testing
agree. We stated clearly in the third paragraph of our discussion that we had made no attempt to assess the scientific validity of the RMCT system. Our concern was with the clinical usefulness of the test. We are aware of reports of good correlations between the RMCT, the RAST and skin tests. In our study, however, the correlations between these tests were poor. We believe that our studies were conducted carefully, with compulsive attention to detail - considerably beyond what might be expected in a busy service laboratory. Furthermore, our experience With the RMCT is not unique. Vijay and colleagues7 stated that, "in spite of painstaking efforts, we were unable to reproduce the technique of rat mast cell degranulation for the detection of human reaginic antibodies, as described by Perelmutter and his colleagues and by others." Perelmutter suggests that negative results for the RMCT with highly atopic serum, as presented in Table III, may be the result of performance of the test in antigen excess. We do not believe that this is likely. First, the results of several tests with the same serum samples yielded widely varying results with no relation to antigen concentration and no suggestion of a doseresponse relation. Second, we used a 10000-fold concentration range of whole, crude ragweed extract (0.0002 to 2% w/v), which is likely within the concentration range of semipurified ragweed antigen used by Perelmutter and Liakopoulou.' Serum used for reproducibility studies was stored for a few weeks, not several weeks. We chose to store the serum at 40C for short intervals to avoid deterioration of the samples by freezing and thawing on the advice of Dr. Perelmutter. However, some studies were done on serum stored at -200C; the results were comparable to those obtained with serum stored at 40C. Contributions to the Correspondence section are welcomed and if considered suitable will be published as space permits. They should be typewritten doubled spaced and, except for case reports, should not exceed 1½ pages in length.
CMA JOURNAL/SEPTEMBER 17, 1977/VOL. 117 569
that for the purpose of performing the reproducibility experiments the serum To the editor: I have several comments was stored at 40C for several weeks. concerning the conclusions reached by I would not advocate such storage conW.B. Chodirker and A. Piotrowska in ditions for this type of experiment since their article entitled "Allergy testing: these, in my experience, have resulted comparison of skin and in vitro tests in similar inconsistent responses. For of allergic reagin" (Can Med Assoc J repeated testing we store serum in small 116: 1254, 1977) as one who has been aliquots at -200C. Chodirker and Piotrowska state that involved with several of the procedures mentioned in the article, particularly the study by Renoux and colleagues3 showed that rat mast cells did not bind the rat mast cell test (RMCT). The authors state that "the RMCT human IgE, but in fact the study was found to be poorly reproducible showed the opposite. Furthermore, and unreliable." While I do not defend Renoux and colleagues used three inthe clinical usefulness of the RMCT dependent techniques to demonstrate as a commercial test to detect imme- binding of* human IgE to rat mast cells. diate hypersensitivity reactions, I be- In addition, Plaut, Lichtenstein and lieve that the conclusions drawn by the Bloch4 actually showed that there was authors do not reflect the scientific a lack of histamine release from rat validity of the test. I have shown pre- mast cells incubated with ragweed alviously that there is a good correlation lergic serum and challenged with ragbetween the RMCT, the radioallerg&- weed antigen, and that this had nothing sorbent test (RAST) and intradermal to do with morphologic changes in rat skin tests.1 A double-blind study was mast cells. Moreover, several investigadone with serum from birch-allergic tors not mentioned in Chodirker and patients. Dr. S.G.O. Johansson of Piotrowska's article have demonstrated Sweden performed RAST analysis and histamine release from rat mast cells..7 Chodirker and Piotrowska imply that sent Dr. K. Ishizaka of Baltimore, Maryland nine samples of serum from I was involved in their study on a both atopic and nonatopic individuals. continuing basis. Although I was inDr. Ishizaka coded the samples and volved initially I cannot be held resent them to me for RMCT determina- sponsible for their experimental contions. An excellent correlation was ditions and conclusions. Furthermore, I believe that articles of this nature found between these in vitro tests. discourage further development of the Table III of the article in the Journal shows that only one of seven serum RMCT, which seems promising for the samples from atopic, highly sensitive, detection of IgE specific for a variety RAST-positive patients gave responses of drugs. It is well recognized that bindin the RMCT, whereas for three of five ing of human IgE by rat mast cells patients with moderate sensitivity the is weaker than binding in the homocyresponses in the RMCT were positive. totropic system; experimental condiThese results are similar to those found tions have to be exact for proper in another of my studies, which showed response in the test. Further work on that a significantly lower allergen con- mast cells from various strains of rats centration is required for serum with a or mice, and investigation into approhigh concentration of IgE antibody and priate storage conditions for these cells, that more allergen is needed for re- may ultimately resolve some of the sponse with lower concentrations of problems encountered with the RMCT. L. PERELMUTTER, PH D antibody.1 It is possible that the reacChief, hematology tions reported in Table III for the highBureau of clinical chemistry and hematology Health protection branch ly sensitive patients were performed in Health and Welfare Canada Ottawa, Ont. antigen excess, when such responses would be inhibited. To the editor: Dr. Perelmutter indicates As to the reproducibility of the that our conclusions do not reflect the RMCT, Chodirker and Piotrowska state scientific validity of the RMCT. We
Allergy testing
agree. We stated clearly in the third paragraph of our discussion that we had made no attempt to assess the scientific validity of the RMCT system. Our concern was with the clinical usefulness of the test. We are aware of reports of good correlations between the RMCT, the RAST and skin tests. In our study, however, the correlations between these tests were poor. We believe that our studies were conducted carefully, with compulsive attention to detail - considerably beyond what might be expected in a busy service laboratory. Furthermore, our experience With the RMCT is not unique. Vijay and colleagues7 stated that, "in spite of painstaking efforts, we were unable to reproduce the technique of rat mast cell degranulation for the detection of human reaginic antibodies, as described by Perelmutter and his colleagues and by others." Perelmutter suggests that negative results for the RMCT with highly atopic serum, as presented in Table III, may be the result of performance of the test in antigen excess. We do not believe that this is likely. First, the results of several tests with the same serum samples yielded widely varying results with no relation to antigen concentration and no suggestion of a doseresponse relation. Second, we used a 10000-fold concentration range of whole, crude ragweed extract (0.0002 to 2% w/v), which is likely within the concentration range of semipurified ragweed antigen used by Perelmutter and Liakopoulou.' Serum used for reproducibility studies was stored for a few weeks, not several weeks. We chose to store the serum at 40C for short intervals to avoid deterioration of the samples by freezing and thawing on the advice of Dr. Perelmutter. However, some studies were done on serum stored at -200C; the results were comparable to those obtained with serum stored at 40C. Contributions to the Correspondence section are welcomed and if considered suitable will be published as space permits. They should be typewritten doubled spaced and, except for case reports, should not exceed 1½ pages in length.
CMA JOURNAL/SEPTEMBER 17, 1977/VOL. 117 569
