Intradermal Tests with Autologous White Blood Cells in Chronic Discoid Lupus Erythematosus, Systemic Lupus Erythematosus and Control Subjects1

INTRADERMAL TESTS WITH AUTOLOGOUS WHITE BLOOD CELLS IN CHRONIC DISCOID LUPUS ERYTHEMATOSUS, SYSTEMIC LUPUS ERYTHEMATOSUS AND CONTROL SUBJECTS * THEODORE A. TROMOVITCH, M.D. AND CYRIL MARCH, M.D.

In the past few years numerous auto-immune

discoid hE., 9 patients with lymphoblastoma or

phenomena have been described in systemic leukemia, 20 patients with atopy, (19 patients with lupus erythematosus (SEE.). Reactions against atopic dermatitis with or without associated cellular components including whole nuclei asthma and allergic rhinitis and 1 patient with allergic rhinitis alone), 20 patients with mis(1—3), nucleoprotein (2, 4), D.N.A. (5—7), histone

(3) as well as cytoplasmic elements (8, 9) have been recorded. These reactions have been shown to be mediated through serum antibodies. Recently another auto-immune phenomenon of a delayed hypersensitivity reaction against intradermally injected white blood cells has been described. Holman (10) found positive reactions to a white cell homogenate injected into patients with S.L.E. Friedman, Bardawil, Merrill and Hanau (11) tested 79 patients to either autologous or homologous whole leukocyte suspensions. They noted

that 16 of 20 patients with systemic lupus

cellaneous dermatologic conditions (psoriasis (1),

lichen planus (1), porphyria (2), alopecia areata (1), lichen simplex chronicus (1), contact derma-

titis (3), male pattern alopecia (9) and polymorphous light eruptions (2). In additiou 10 normal volunteers, having no known disease, served as a control group. The diagnosis in each case was arrived at from both the clinical picture

presented at the time of testing and a careful review of the patient's chart. Patients with L.E. were classified as having the chronic discoid, subacute, or acute varieties on the basis of the skin lesions, history, laboratory findings and the course of the disease.

The term C.D.L.E. in this study is used to

erythematosus and 2 of 7 patients with designate patients who displayed the classical rheumatoid arthritis manifested positive reactions. In their control group only 1 of 52 patients

and purely cutaneous form of C.D.L.E., whether localized (30 patients) or generalized (1 patient)

and who had a negative hE. cell preparation and no consistent multisystem findings which have published results similar to those obtained would suggest a diagnosis of systemic L.E. It by Friedman et.al. (11) in S.L.E. The following should be noted, however, that S of the 31 patients with C.D.L.E. did complain of arthralgias manifested a positive test. Recently Bennett and Holley (12) using homologous leukoeytes

study was undertaken to evaluate the reaction to intradermally injected autologous white blood

and that 4 had elevated sedimentation rates.

cells in patients having chronic discoid lupus Preparation of Test Materials erythematosus (C.D.L.E.), other cutaneous The materials for testing consisted of autoldiseases and in healthy control subjects. ogous 1) leukocyte suspension, 2) red blood cell suspension, and 3) plasma.* The method reported

MATERIALS AND METHODS

by Friedman et at. (11) was used as follows:

Clinical Material

The subjects tested included 20 patients with systemic L.E., 31 patients with typical chronic * From the Department of Dermatology of the

New York University Schools of Medicine and the

Skin and Cancer Unit of University Hospital,

New York, N. Y. This study was supported by a grant from the United States Public Health Service (E-1361-(C5) and was done during the tenure of a special fellow-

ship from the Allergy Foundation of America through funds provided by Burroughs Wellcome Co. (U.S.A.), Inc.

approximately 27.5 cc of venous blood are drawn into a 30 ml glass syringe containing 10 mgm of

heparin and 400 mg of dextrant (2 cc of a 20% aqueous autoclaved solution). The needle is then occluded by insertion into an autoelaved cork and the syringe placed in the refrigerator at 4° C in a vertical position resting on the plunger. After 15 to 60 minutes in this position two distinct layers appear—a lower layer of sedimented R.B.C. and * Test materials also contained 0.03 mg heparin and 1.3mg dextran per 0.1 ml plasma.

t Liquanemin Sodium, Organon Inc., Orange, * Pharmachem Corp. (Formerly R. K. Laros Co.) Bethlehem, Pa.

Presented at the twenty-second Annual N. J.

Meeting of The Society for Investigative Dermatology, Inc., New York, N. Y. June 29, 1961.

345

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an upper straw-colored layer of leukocytes and platelets suspended in plasma. The upper strawcolored layer is expressed into a centrifuge tube by maintaining the syringe in vertical position, applying pressure to the plunger and forcing the fluid through the needle bent at a right angle. Ten to 15 cc of straw-colored material usually obtained by this procedure is then centrifuged at approximately 2000 rpm for ten minutes in order to separate the leukocytes. The plasma is pipetted off until only a 1 cc volume remains in the tube. The remaining plasma is then thoroughly mixed

mononuclear cells with a small number of polymorphonuclear cells and histioeytes. No vascular necrosis was seen. Bennett and Holley (12) who have studied the histology of these reactions more extensively stressed the mixture of mononuclear and polymorphonuelear cells in the perivascular

infiltrate. In addition, by means of Masson's triehrome and phosphotungstic aeid-hematoxylin stains, they demonstrated a fibrinoid transudate around blood vessels. RESULTS

with the leukocyte button to produce a homogAll tests performed with red blood cells and enous leukocyte suspension. This constitutes the plasma were negative except in one subject who leukocyte fraction. The cell free plasma is used as the second test fraction and the sedimented red blood cells which remain in the syringe constitute

had seasonal allergic rhinitis at the time the skin tests were done and who gave a positive

reaction to all three test materials. Three months later he was retested and showed a positive test fraction showed white blood cells of both granulo- to only the white blood cells. All other reactions cytic and lymphocytic series with a normal differ- observed by us were to the white cells; therefore,

the third test fraction. In our studies, smears of the white blood cell

ential count and the presence of platelets. The "positive test" as used hereafter refers to the "contamination ratio" (number of W.B.C./num- skin test with autologous white blood cells only. ber of R.B.C.) varied from 1:1 to 1:10. The leuko- Table 1 gives a summary of results obtained by cyte fraction showed considerable variation in its W.B.C. count per mm3, as was to be expected, testing with autologous white blood cells. Group I. Systemic L.E. There were 20 patients depending upon the W.B.C. count of the test subject; the average count in this fraction was in this group. Seventeen had acute systemic L.E. 65,000 W.B.C./mm3 and the range was 19,000 to at the time of the skin test. Sixteen of these 17 236,000 W.B.C./mm3. The amount of W.B.C. cases had a positive L.E. cell preparation. Skin used in the skin test showed no correlation with lesions were present in 12 of these 17 patients. the results obtained (see "Results"). Five patients had no cutaneous lesions. The lesions in 10 patients consisted of a butterfly Method of Skin Testing erythema sometimes associated with an Intradermal testing was done on the flexor ephemeral patchy erythematous macular erupsurface of the forearm with 0.1 cc of each of the tion covered by fine scaling. Two patients had three test fractions. Tuberculin syringes and lesions suggestive of a generalized discoid type; 2125 needles were used. The injections were made

immediately following preparation of the test fractions and the sites were examined twenty-four

hours later. A test was considered positive if an area of induration greater than 10 mm associated with a surrounding erythema was present. The average positive test showed 16.8 mm of induration and

20.1 mm of erythema. A test was considered negative if only erythema or induration less than

10 mm was present. The average negative test measured 6 mm of combined induration and erythema. Positive reactions began to appear within 12—18

hours after injection, reached a maximum at 24 hours and at 48 hours were definitely diminishing. At 72—96 hours the reaction had completely dis-

appeared. Histologically these positive white blood cell test sites showed a banal perivascular

inflammatory infiltrate consisting mainly of

TABLE 1 Results of intradermal W.B.C. skin tests Number coup

I. Systemic L.E II. Chronic Discoid L.E III. Normal IV. LymphoblastomaLeukemia V. Atopic disease.. .. VI. Miscellaneous

of Subjects Tested

Number Positive to Skin

Test

Number Negative to Skin

Test

20

19

31 10

10

21

0

10

9

4 5

15

3t

17

20 20

1*

5

* This patient was in remission at the time of the W.B.C. skin test. t These 3 patients had a history of arthralgia.

TESTS WITH LEUKOCYTES IN LUPUS ERYTHEMATOSUS

however, no definite atrophy was present. There

347

Group VI. Miscellaneous dermatoses. A positive

was no correlation between a positive white white blood cell skin test was observed in 3 out blood cell skin test and the presence of skin of the 20 cases with miscellaneous dermatologic lesions or any particular type of skin lesions. conditions (2 patients with male pattern alopecia All 17 patients responded with a positive white and one with a polymorphous light eruption.) blood cell skin test. Two patients were considered

to have systemic L.E. of the subacute variety; they had positive L.E. cell preparations but were

All 3, in addition to their dermatologic problem, had arthralgias. An L.E. cell preparation could be done only on one of these 3 patients and was

not acutely ill. Both showed positive W.B.C. negative. skin tests. One subject had a positive L.E. cell DIsCUssION preparation 3 months earlier but at the time of A hypersensitive reaction to intradermally skin testing was in apparent clinical remission and on no medication. This patient had a negative injected leukocyte suspension appears well white blood cell skin test. Another patient in this established in S.L.E. Friedman et al. (11) using series who showed a positive W.B.C. skin test autologous and homologous leukocytes noted while she was in an acute phase went into remis- positive findings in 16 of 20 S.L.E. patients. sion during chioroquine therapy. Subsequently Bennett and Holley (12), using a modified while her disease was in remission and at a time technique, recently reported positive skin tests when she no longer was taking chloroquine, her in 15 of 17 patients with S.L.E. In our studies, 19 of 20 patients with S.L.E. manifested a white cell skin test was negative. Group II. C.D.L.E. The 31 patients in this positive test. It is interesting that the incidence of positive group had the typical lesions and clinical course of C.D.L.E. with no evidence of multi-system skin tests in this series paralleled the severity involvement which would permit a diagnosis of and acuteness of the disease process both in S.L.E. L.E. cell preparations were performed on terms of acute symptoms and in the severity of 26 of these 31 patients and were negative. Ten the form of L.E. All S.L.E. patients in an active of these 31 patients had a positive white blood phase were positive whereas patients who were in skin test. L.E. cell preparations were performed remission and taking no medication gave negaon 9 of these 10 positive reactors and were all tive tests. No direct relationship between positive skin test and organ systems involved by negative. Group III. Normal control subjects. The 10 disease could be found. The nature of the antigen which is responsible personnel working at the Skin and Cancer Unit for the delayed intradermal reaction to autologous of University Hospital. No positive white blood W.B.C. has not yet been defined. Investigations normal control subjects in this group were healthy

cell skin tests were elicited in any of these control to characterize the antigen are currently in progress. subjects. The finding of 10 positive skin tests in 31 Group IV. Leukemia—lymphoblastoma. Nine

patients in this group were diagnosed as having cases of classical chronic discoid L.E. may be leukemia or lymphoblastoma. Four of these interpreted as another indication that C.D.L.E. gave positive skin tests—2 had chronic lymphatic even when not accompanied by signs of systemic leukemia, 1 had Hodgkin's sarcoma, and 1 had involvement is, at least in some cases, part of stem cell leukemia. Negative white blood cell the same disease complex as S.L.E. Bennett skin tests were observed in one patient each with et al. (13) found 2 positive responses in 14 patients Brill-Symmer's disease, Hodgkin's disease, malig— with C.D.L.E. The difference between our results nant lymphoma, monocytic leukemia and acute may be due to their use of a slightly different method for preparing the W.B.C fraction, to a myelogenous leukemia.

Group V. Atopic dermatitis and allergic difference in criteria for classifying patients in the

rhinitis. Of the 20 patients in this group, 4 out of 19 having atopic dermatitis alone or in combination with hay fever or asthma manifested positive skin tests. In addition, one patient with hay fever alone also showed a positive white blood cell skin test.

various groups or to statistical factors inherent in

the small number of patients tested. Whether a positive W.B.C. skin test in C.D.L.E. indicates

those patients who are particularly liable to develop the subacute or acute form of L.E. later on remains to be seen through long terms follow-

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up. No correlation could be found in C.D.L.E.

patients with atopy also is noteworthy. Atopies

patients between a positive white blood cell characteristically tend to have multiple allergic skin test and the duration or extent of disease, responses of the immediate type to common presence of arthralgias, elevated sedimentation protein allergens. Whether these reactions against rate, depressed or elevated white cell count, white cells are due to inhaled or ingested protein differential count, hemoglobin, or urinary find- antigens phagoeytized by the white cells and ings. This appears to preclude the possibility slowly released at the test site by the injected that leukopenia in S.L.E. is due to the same leukoeytes or whether persons with a hereditary factors which are responsible for the positive atopie background tend to develop delayed skin reaction to W.B.C. hypersensitivity to W.E.C. constituents or other

The strong possibility that this reaction is autologous materials cannot be answered at mediated through a delayed type of hypersensi- this time. tivity mechanism is one of the most provocative The three positive responses occurring in conaspects of the W.B.C. skin test. Reports con- trol patients having arthralgias associated with cerning the immunology of L. E. have in the male pattern alopeeia and polymorphous light past focused on serum antibodies. These have eruption cannot be readily explained at present. been demonstrated against whole nuclei (1—3), The fact that these 3 patients also had arthralgias nueleoprotein (2, 4), D.N.A. (5—7), histone (3) perhaps is of significance since Friedman et at. and eytoplasmie elements (8, 9); however, there (11) reported positive tests in 2 of 7 patients with

is no general agreement as to the importance of these antibodies in the pathogenesis of L.E

rheumatoid arthritis. Our 3 patients will be

followed clinically and with appropriate laboraIt is tempting to speculate that the pathogenefie tory tests. changes seen in L.E. are the result of an autoOur results make it obvious that the W.B.C. sensitivity reaction mediated through cellular skin test is not pathognomonie nor specific for

and serum factors. This would explain why lupus erythematosus. It remains to be seen during the first few months infants born of whether a more specific skin test for L.E. can be mothers with acute L.E. do not develop manifes developed through the use of purified white tations of L.E. (14, 15) even though they may blood cell fractions. The significance of the have positive L.E. cell preparations. Apparently positive W.E.C. skin tests in eases of leukemia, a sufficient quantity of serum antibodies hut an lymphoblastoma and atopy also requires further insufficient number of white blood cells nae investigation. SUMMARY through the placental barrier. The finding of positive skin tests in lymphoDelayed cutaneous hypersensitivity to autoloblastoma and leukemia patients presents another gous leukoeytes was shown in 17 of 17 patients interesting problem. Others have reported posi with acute systemic L.E. 2 of 2 patients with tive skin tests to lysed W.B.C. in acute leukemia subacute L.E. and in 10 of 31 patients with (16), and to cancer tissue in malignant melanoma. chronic discoid L.E. A negative test result was (17) and various carcinomas (18). It is particu obtained in 2 patients with S.L.E. in remission.

larly noteworthy that the positive skin tests

Positive tests were elicited also in three other occurred in our patients with leukemia or lyni- distinct groups: patients with leukemia or phoblastoma involving stem cells or lymph lymphoma, patients with atopie dermatitis and

nodes. Does the positive test represent the allergic rhinitis and patients with other

host's response against antigens in the malignant dermatoses. cells or is this a phenomenon in which malignant, Ten healthy control subjects gave negative yet immunologically competent cells are reacting test results.

against the host's normal cellular constituents Evidence to date suggests that the skin reacin a manner similar to the reaction which pro- tion to autologous W.B.C. represents a delayed duces "runt disease?" Further studies utilizing hypersensitivity response against leukoeytes passive transfer technic might produce a better mediated through immunologically competent understanding of the nature of these reactions in cells rather than serum antibody. The possible patients with leukemia and lyniphoblastoma. significance of positive skin tests to autologous The finding of positive skin tests in 5 of 20 W.B.C. in L.E. and other diseases is discussed.

TESTS WITH LEIJKOCYTES IN LIJPIJS EHYTHEMATOSIJS

H. C.: Anticytoplasmic factors in the sera of patients with systemic lupus erythematosus and certain chronic diseases. Arthritis

ACKNOWLEDGMENTS

The authors wish to express their gratitude particularly to Drs. R. L. Baer, L. C. Harber, A. Hyman, P. Miescher, N. Orentreich, N. Rothfield and L. Weiner of New York University Medical Center and to other staff members of the

Skin and Cancer Unit who encouraged this study. Credit for technical assistance is given to Miss M. Gillman. REFERENCES 1. MIE5CIJER, P. AND FAIJCONNET, M.: L'Absorp-

tion du facteur "L. E." par desnoyaux cellulaires isolds. Experientia, 10: 252-54,

349

Rheum., 3: 1—15, 1960.

10. HOLMAN, H. as cited by BENNETT, J.

C. AND

HOLLET, H. L.: Intradermal hypersensi-

tivity in systemic lupus erythematosus.

Arthritis Rheum., 4: 64—73, 1961. 11. FRIEDMAN, E., BAEDAWIL, W, MERRILL, J.

HANAU, C.: "Delayed" cutaneous hypersensitivity to leukocytes in dissemiAND

nated lupus erythematosus. N. Engl. J. Med., 262: 486-91, 1960.

12. BENNETT, J. C. AND HOLLET, H. L.: Intradermal hypersensitivity in systemic lupus erythematosus. Arthritis Rheum., 4: 64—73, 1961.

13. BENNETT, J. C., O5MENT, L. S. AND HOLLEv,

H. L.: Immunological manifestations in a group of patients with discoid lupus erythematosus. Presented at the Hahnemann Hospital, Symposium on inflammatory and connective tissue diseases, Philadelphia,

1954.

2. HOLMAN, H. AND KUNKEL, H.: Affinity

between the lupus erythematosus factor

and cell nuclei and nucleoprotein. Science, 126: 162-63, 1957.

1960. (To be published.)

3. HOLMAN, H., DIEcHEE, H. AND Ronnixs,

C.: Antinuclear antibodies in lupus 14. BRIDGE, R. G. AND FOLEy, F. E.: Placental transmission of the lupus erythematosus erythematosus. In "Symposium on Hyperfactor. Amer. J. Med. Sci., 227: 1, 1954. sensitivity", pgs. 361—9. Boston, Little, Brown & Co., PubI., 1959. 15. BEELTNE, G. M., SHORT, I. A. AND VIcKEE5, C. F.: Placental transmission of the L. E. 4. SCALETAR, R., MARCUS, D. M., SIM0NT0N, L. A. W.

AND MU5CHEL, L. H.: The nucleoprotein complement fixation test in the diagnosis of

5.

systemic lupus erythematosus. N. Engl. J. Med., 263: 226-29, 1960. MIEsCHEE, P. AND STEA55LE, R.: New serological methods for the detection of the L.

factor. Lancet, 2: 5, 1957.

16. BERNARD, J., GEABAE, P., SELIGMANN, M. AND BADILLET, M.: Intradermo-reactions a

des extracts de leucocytes normaux et de leucocytes leucemiques chez des sujets atteints de leucemie aigue. Bull. Soc. Med.

E. factor. Vox Sang. Basel, 2: 283-87, 1957. 6. CEPELLINI, R., POLLI, E. AND CELADA, F.:

Hop., Paris, 70: 1169, 1954. Cited in Walford,

R. L.: Leukocyte antigens and antibodies p. 42. New York and London, Grune & Stratton, 1960.

A DNA reacting factor in serum of a patient

with lupus erythematosus diffusus. Proc. Soc. Exp. Biol. Med., 96: 572, 1957.

7. DEICHEE, R., HOLMAN, H. AND KUNKEL, H.:

The precipitin reaction between DNA and a serum factor in systemic lupus erythematosus. J. Exp. Med., 109: 97-114, 1959. 8. AauEasoc, G. L.: Antibodies against nuclear

and cytoplasmic cell constituents in systemic lupus erythematosus and other disease. Brit. J. Exp. Path., 40: 209, 1959.

17. MAKARI, J. H.: Demonstration of antibodies to cancer polysaccharides and polysaccharides and polysaccharide-antibody com-

plexes by an intradermal reaction. J. Amer. Geriat. Soc., 8: 4-15 1960.

18. MAKARI, J. 0.: Recent studies in the immunology of cancer. V. Detection of tumors in man

by the skin testing of polysaccharideantibody complexes. J. Amer. Geriat. Soc.,

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9. DETcILEE, H. R., HOLMAN, H. R. AND KUNKEL,

1960.

DISCUSSION DE. DONALD

When we

J. BLANEv (Cincinnati, Ohio):

read the initial report by Freidman

at., we were impressed, and we did some intradermal leukocyte tests, as did the presenter. We came up with similar results, et

similar

some false-positives.

If it would prove applicable, obviously it would be very simple to have some lyophilized nucleoprotein, just as we do tuberculin, and do skin tests to screen for S.L.E. So

we tried it,

results,

and

we came up with

similar

that is, similar to testing with white

Since the antibodies that are present in the blood cells. We have seen false-positives just as circulating plasma in S.L.E., the gamma globulin

antibodies, are nonspecific insofar as they will react with nucleo-protein from any source, it

seemed

logical that the delayed type antibodies

might also be nonspecific and we could use

regularly and also false-negatives in some cases of L.E. A generalized swelling rather than discrete induration sometimes makes interpretation of the test difficult. Our series is not as extensive as the presenter's,

nucleoprotein from a more convenient source, but I would say that in our hands the skin test has not as yet been useful diagnostically; but calf thymus, as an intradermal skin test.

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THE JOURNAL OF INVESTIGATIVE DERMATOLOGY

it is interesting and perhaps explains some manifestations of L.E. that are not explained by

as positive reactions, which occur for reasons not presently understood.

serum antibodies. DR. MARION B. SULZBERGER (New York,

In viewing this data it is interesting to note that these positive reactions tend to fall into

New York): First of all, I want to say that this

separate diagnostic groups and arc not randomly distributed among all the disease states tested. I want to compliment Dr. Tromovitch and Dr. This was not completely predictable at the start March and coworkers on the great conservatism of this study. is a very well presented piece of careful work and

which they have used in expressing and drawing conclusions as to the mechanism and what the results mean. Then, I would like to hear tbeir comments about the difference between this and the usual delayed form of response. The early disappearance of this reaction they have noted at a time when the usual "delayed" form of skin response, the tuberculin or, 48-hour-type of response is just about reaching its maximum and beyond which it then usually continues, points to some difference here.

I am very much interested in the skin test reactions to nuclcoprotein, particularly the source of the nucleoprotcin because Bennett and Holley (Ref. 12) have used one type of nucleo-

protein and have not obtained positive skin tests.

In a patient of Dr. Peter Miescher whole nuclei from trophoblasts gave a negative skin test even though she was positive to the white blood cell skin test done at the same time. In answer to Dr. Sulzbcrgcr's question regarding the difference between this reaction pattern

The second point I would like to make is that and that of the tuberculin type, the former if my figures arc correct (and I added up quickly probably fits best with what is called the inter-

while listening to Dr. Tromovitch and looking mediate type or the so-called bacterial type of at the slide), and if you divide the series in the hypersensitivity. This 24 hour type reaction is following way: the lupus erythcmatosus acutus seen as a response to bacterial antigens when used or systemic lupus erythematosus cases in one to skin test patients sensitive to them. group and all the others as controls (that is the As far as the validity of the test in each

blood dyscrasias, the atopic dermatitis, the category is concerned, I agree that the data miscellaneous dermatoses and the normals) in presented is insufficient from which to draw

the other group, one finds that twenty-two out of the ninety "controls" arc reacting to this skin test with white cells—i.e. just about one-quarter of all of the "controls" react. To my mind this points up the fact that one needs a great many more "normals" as controls here to sec if the "normals" arc really non-reactors in regard to this test and whether this 25% incidence of reactors is not just the incidence of positive reactions to leukocytes which will occur in a general population which excludes acute

definite conclusions. I think, however, that the finding of positive skin tests in certain groups is consistent with present knowledge. For example, in the leukemia group one might expect to obtain positive tests in the light of all the previous work on other types of cancer where immunologic responses have been shown against the neoplastic tissue.

I believe it was Dr. Fisher who asked

about immediate responses. We did not tabulate immediate responses because many times when lupus erythematosus. DR. ALEXANDER A. FISHER (Woodside, Long injecting simply serum, particularly into atopic patients, a transient reaction would be seen. Island, New York): I would like to know how We followed the delayed response which began

many immediate responses you had in your series of "specific" cases or "unspecific" cases. How did you decide that it is a delayed type

about 12 hours following the injection of the

test material. In regard to biopsies, the microscopic picture of reaction as we know eczcmatous or tuberculin? is not impressive, showing some perivascular Why couldn't it be an Arthus's phenomenon? Did you make any biopsy studies to decide that inflammatory infiltrate of lymphocytes, a few histiocytes and a few polys. question. DR. THEODORE A. TR0M0vITcH (in closing):

Bennett and Holley reported that with

I with to thank the discussors for their comments.

trichrome and phosphotungstic acid stains they could demonstrate a transudate around blood vessels which had a fibrinoid appearance.

In regard to Dr. Blaney's question, I prefer to refer to these reactions not as false positives but

THE JOURNAL OF INVESTIGATIVE DERMATOLOGY

94

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