TOXICOLOGYAND APPLIEDPHARMACOLOGY)SUPPLEMENTNO. 3, 30--44 

### (1969)



### Evaluation of Cosmetics for IrritancyZ

ALBERTM. KLmMAlq DepartmentofDermatology, UniversityoflaennsylvaniaSchoolofMedieOte, Philadelphia,Pennsylvania29104
"When description gives way to measurement, calculation replaces debate."
STEVENS

As with hostility, every one instantly recognizes what is meant by irritancy; nonetheless, our knowledge of these parlous subjects is more descriptive than definitive. When we are obliged to provide exact definitions, modes of action, and especially appraisals of magnitude, we soon become irritated by ambiguities and vacuities. We have in our various roles as toxicologists, lawyers, biologists, physicians, and guarantors of the public weal a keen subjective sense that irritants are harmful, but objective criteria for estimating the hazard are hard to come by and in any case controversial. Despite these limitations I should like to make it clear at the outset that the general record ofthe cosmetic industry has been veryfavorable as regards cutaneous toxicity, attesting at least to prudence and conservativeness evenin the absence of sophisticated procedures oftesting. I do not regard this as an emergency session to straighten out a crooked enterprise. Yet it is quite clear that various forces are converging which will necessitate the development of a more reliable and uniform methodology. The distinction between drugs and cosmetics, always more casual than correct, must break down under critical scrutiny. Surely the thrust ofmodern technologyis toward greater "activity". Enterprising houses are increasingly interested in improving the effectiveness of their products through the opportunities provided by the dazzling inventiveness of the synthetic chemists. Can anyone seriously defend the position that preparations to combat acne, dandruff, infections, and excessive perspiration are pharmacologically inactive? Originally cosmetics were designed to conceal or decorate; now we must acknowledge that they are often intended to ameliorate or prevent cutaneous abnormalities--dare
one use the terrifyingword "disease" ? In the near future we can expectthat cosmetics, at least those which influence cutaneous functions, will have to meet the same standards as drugs. This ofcourse means governmental surveillance, and I, for one, think the net effect will be salutary, though probably painful. It is no longer adequate to depend upon sales as an index of safety. Moreover, the consumer population is more disposed toward damaging legal actions, oftenwithout realjustification. More exacting regulations for safety testing will, in the end, protect rather than harass the manufacturers.

1Thisstudywasconducted underthesponsorshipoftheCommissionon Cutaneous Diseasesofthe Armed Forces Epidemiological Board. 30COSMETIC IRRITANCY 31

Whereas the primarypurpose ofa drug isto cure, cosmetics are supposed to delight; accordingly, there is practically no tolerance for harmful reactions no matter how trivial or transient. The incidence ofcutaneous toxicitymust perforce be so low as to engender insomnia and onyehophagia in those who are ultimately responsible for approvingthe sale oftheproduct. For example,ifonly one in a thousand users became irritated, there would be for every million units sold, a thousand complaints, enough to frighten the mamffacturer and alienate the consumer. One cannot too strongly emphasize the peculiarly stringent requirements which cosmetics must meet as regards irritativeness. In fact the central problem in testing cosmetics is precisely that one is generally dealing with products of a very low order of toxicity. Accordingly we shall need particularly sensitive techniques ofappraisal

The burden of this discussion is on local irritativeness, not systemic effects. It is my intent to sketch out the principles, strategies, and tactics which may be utilized in measuring irritativeness. Although a formal consensus does not yet exist, I rather expect that the ideas presented, none of them revolutionary or novel, will accord with conclusions that experiencedpersons generally hold. Because we arejust beginning to reduceprinciples to practice, it will be necessaryto stress strategies rather than dictate tactics. The following script is to be regarded as a framework for the eventual establishment ofstandards not as a straitjacket to force conformityto a particular method. 

### GeneralPrh2ciples

Irritants are toxic substances which directlydamage tissues. This is to some extent a definition by exclusion, encompassing all adverse 

### local effects which are nonallergic. Simple as it is, the subject is confounded by curious notions and an archaic nomenclature. For example, it is the convention to place the adjective "primary" before the term irritant so that one speaks about "primary irritants." The strict semanticist will then wish to know how these differfrom "secondary" or even "tertiary" irritants. It has been suggested that "primary" irritants irritate on the first exposure and 

### secondary irritants after multiple exposures, but this leads to the complication, if not contradiction, that a substance may be both a primary and a secondary irritant depending on concentrat~o~ alone (Shelanski, 1951). The term primary irritant is a misconception; "primary" is redundant, and its permanent expulsion will encourage clearer description and more rigorous thinking.

Far more serious is the tradition, glaringlyvisible in teaching texts, ofdividing substances into two classes, irritants and nonirritants, implying that oneneed onlyconsult
an authoritative list to decide whether a substance is likely to be harmful or not. This is not merely an oversimplication, it is specious in the extreme. No substance is 

### always safe,nor

### ahvays harmful. The distinctionwe are afteris quantitative not qualitative.

### All substances arepotential irritants. They may vastly differin degree ofirritativeness, but recognizingthe principle has the beneficial effectofcompelling us to think in forms of "how different"? We can answer this only by a measurement of some kind, and such measurement is obtainable only by procedures which are susceptible to quantitation. The goal then becomes a rigorously definedmethodology which will yield figures that
can be treated statistically.

My insistence that all substances should be regarded as potentially irritating is by
no means an academic doctrine. If we take the instance of water, the "harmless"

### 32 KLIGMAN

substancepat" excellence, 

### we can nevertheless envision situations in which the effectis harmful. For example, keepingthe skinwet under occlusive dressings or byprolonged immersion can readily lead to closure of the sweat pores provoking sweat retention lesions ofmiliaria. Or, the enormous stimulusto bacterial and fungalgrowth furnished by wetness and hydration of the horny layer may elicit pyoderma or moniliasis. Admittedly the mechanism of injury is indirect, but the consequences are harmful nonetheless. In any case, these peculiar reactions will have to be taken into account when interpreting closed patch tests under which hydration is unavoidable. In hot weather miliaria is easily mistaken for an irritant reaction. So for that matter are staphylococcalinfection andmoniliasis.Each ofthesefalsepositiveexpressions has led
me into error, indeed with such frequency that one ought constantly to maintain a high index of suspicion. Still another erroneous notion is that individual differencesin susceptibility to irritation are not thought to be very great. It is usually taught that only a proportion of the population is likelyto become allergicto a sensitizerwhereas all persons will react to an irritant. This is a falseconceptionwhichcomes aboutbecause ofthe unconscious tendency to think that irritants are strong and allergens mild. However, one could argue the opposite case. For example, with potent allergens such asp-phenylenediamine or tetrachlorosalicylanilide all adequately exposed persons will become sensitized. By contrast with weak irritants such as 1~o hexachlorophene in mineral oil or Hydrophilic Ointment U.S.P. (containing 1 ~ sodium lauryl sulfate), no amount of exposure is ever likely to prove irritating to more than a small proportion. In this instance, the differencebetweenirritants and sensitizersisnot quantitative but qualitative. The frequency of reactions cannot help one decide whether one is dealing with
one rather than the other, not to mention the complication that many substances are both.

Even experienced persons sometimes do not appreciate the vast inherent differences in susceptibility to irritation. This is because data have usually not been collected under sufficientlyrigorous conditionsto enable oneto plotfrequencydistributions and obtain an ideaofdispersion. Everyonerecognizes that somepersons react more swiftly
or more intensely than others, but adequate characterization of subject variability requires some kind of dosage response data. For detergents, Justice et al. 

### (1961), in
arm immersiontests and Kligmanand Wooding(1967), usingpatch tests with a variety ofirritants, have demonstrated that the log normal probability plot is, in fact, linear. The slope may be shaIlow and the dispersion great, indicating marked individual variation. To emphasize this point I have collected some new data for the express purpose ofpresenting them in theform ofa frequencydistributionplot. It should be noted that the data are probably more reliable than usual because the subjects were of the
same age and race (20- to 25-year-old, white males), all the testing was done at the
same time, and one person applied all the patches according to a rigidroutine. A 20 aqueous solution of aluminum sulfate was applied under continuous occlusion for 12 days to a forearm site of 100 males. The dressing was changed every other day, the solution being reapplied to the same site by our standard technique (Kligman and Wooding, 1967). For each subject the end point was the number ofdaysjust required to produce an erythema, after which there was no further exposure. Figure 1 is a frequency distribution bar plot based on percentage ofpersons reacting at 2, 4, 6, 8, 10,COSMETIC IRRITANCY 

### 33

or 12 days. Naked-eyeinspectionis sufficientto establish that thepattern more or less approximates a normal, bell-shaped distribution curve although there is some skewness. Clearly, individuals differwithrespect to the amount of exposure that will cause irritation, in somein as little as 2 days and in others 12. The majorityreact by 6 days. Failure to appreciate the range overwhich the irritancy data maybe spread promotes misinterpretations. This problem arises, for example, when one wishes to establish a nonirritatingconcentrationfor allergicpatch testing. Large numbers of subjects must be studiedto establish safelevels. Supposethat only5 ~ react to a concentration which isharmlessfor therest ofthe population. One mightpatch test 25 subjects and findno reactors at this concentration. In future patch tests with this "safe" concentration, reactors would be deemed allergic. There are instances in which patients werejudged

30

~o

I0 

## 7 2ox At2 (so,)~

2 4 6 8 I0 12 Days 

### Fro. 1. Frequency distributionofthe days requiredtojust produce redness in 100males patchtested continuously with20~ aluminum sulfate for 12days.

to be contact-sensitizedto propylene glycol and ammonit~mpersulfate solelybecause they reacted to concentrations which are 

### usually harmless (Warshaw and Herrmann, 1951; Calnan and Shuster, 1963).

The most thorough quantitative study of reactions to a variety of irritants has recently been published as a doctoral dissertation by BjSrnberg (1968) in Sweden, Amongmanyvaluable results was thefindingthat individualscannotbe characterized
as being more or less generally susceptible to irritation. High susceptibility to one irritant is poorly correlated with susceptibilityto another. Each irritantis a case unto itselfand cannot serve to locate individuals at unusuallyhigh risk ofirritation. Thus pro-employmentpatch testingisprobably irrational.

The universe ofchemicals is multitudinous and multifarious. It wouldbe decidedly helpfulifwehad a better understandingofthemode ofaction ofirritants, i.e., the biochemical and pharmacologic events which lead to an inflammatoryresponse. Unfortunately our ignorance of this subject is profound except for the obvious example of34 KLIGMAN

that class of extremely toxic substances which physically degrade the structural components and macromolecules of cells, sometimes lumped together under the term of caustics. Strong acids and bases will hydrolyzeor coagulate proteins, rendering the cell lifeless. Coming down the scale, anionic surfactants and even urea will denature proteins and arrest metabolic function. Dimethyl sulfoxide makes cell membranes so permeable that the vital disequilibrium between the external and internal milieu cannot be maintained. Lysosome labilizers, such as vitamin A, probably excite damage through the release of proteases and acid hydrolases, a more subtle mode of action. Some chemicals provoke urticarial reactions by releasing histamine fi'om mast cells. Other possible effects include enzyme inhibition, complexing of DNA and RNA, and mitotic inhibition. However, as regards the mode of action ofmost mild irritants, we are mainly in the dark. This is a basic biological problem which has yet to be vigorously attacked. That irritants cause very different types of damage becomes obvious when the injured tissue is examined microscopically. Figures 2A to 2I illustrate the variability of the histologic injury. Still another problem concerns the clinical criteria by which contact allergy and irritancy can be differentiated. The classical signs ofdermatitis are chieflyredness and swelling, sometimes accompanied by oozing and blistering. However, these manifestations are stereotyped, common reactions to injury. Though the experienced observer
can often grossly distinguish irritation from sensitization when the reactions are severe, this is definitely not true of mild responses which may consist only of redness and slight swelling. Clinical powers fail utterly. This difl]cultyis not simply something for scholars to worry about. With the Draize-Shelanski repeated insult test I have too often been mistaken in diagnosing irritancy reactions as allergic and vice versa on the



# ~.,,~o ;'4" "2

i~i~lll~ r ;, 9 ~ ~.t.~' , _,-, . ,,,,,- . "l",'~Ik3W" ~

, ~"" ,. ~ .~ " t*-~ ,'~".~"r " ~. :~,

FIG. 2. Variations in response of human cutaneous tissue to different irritants. Hematoxylin and eosin. (A) Cantharidin, intraepidermal blister; the primary event, before cellular damage, is a loss of cohesion between the cells so that they simply fall apart and float freely in the cavity. •COSMETICIRRITANCY 35

Fio. 2 (B) Thickened, hyperplastic epidermis, covered with a crust of killed cells, 4 days after application of a cationic detergent, x I 11.



### ~7~37;;

... . / .

FIG. 2 (C) Follicular abscess following application of high concentration of a corticosteroid, x45.36 KLIGMAN

FIG. 2 (D) Infiltration of leukocytes around intraepidermal sweat duct following application of aluminum sulfate. :<360.

2(X)

F1o. 2 (E) Subcorneal blister with toxicepidermal changes due to kerosene, x 129.COSMETICIRR[TANCY 37

...... ~ ' ~" ~ -41- "~ - ~

Fzo, 2 (F) Subepidermal blister due to sodium lauryl sulfate. The epidermis is surprisingly normal.
• 159,

FIo. 2 (G) Intercellular edema induced by vitamin A acid, a lysosorne]abilizer. The cells are forced apart by fluid, accentuating theprickles. •38 KLIGMAN

"2(H) ' " ..,V ~" 

# N "O 

# III ,"

Fro. 2(H) Follicular abscess due to croton oil. x I t I.

~~,~ . ~'

'S
! ) Nt~

Fro. 2 (I) Completedestruction ofepidermis and necrosis ofdermis produced by a caustic application ofsodiumhydroxide, x219.
final challenge patch test. The only recourse I know of for resolving this issue confidently is biopsy. A 3 mm punch will do. The histologic pattern ofthe allergic reaction is diagnostic, beginning with perivenular accumulations of mononuclear cells accompanied by exocytoses of lymphocytes and spongiosis.COSMETIC IRRITANCY 39 

### So'ategiesfor Appraising b'ritants

While it is obligatory to use human skin for final evaluation, one should not on that account discount the usefulness of animal bioassayin preliminary screening. As a rule, the depilated skin ofrabbits and guinea pigs is more easily irritated, but this can hardly justify a surprisingly widespread anthropocentric bias against animal tests. On the contrary, careful comparative study on both human and animal skin shows quite clearly that the rank order ofirritancy may be much the same (Justice 

### et al., 1961 ).

It must be emphasized that irritancy testing is afflicted with numerous variables whose influence must be minimized. These are too easily disregarded except by those who have suffered the torment of trying to obtain reproducible results. Cutaneous reactivity is influenced by body region (the scalp and face are comparatively resistant), age (children are more easily irritated), sex ([imited evidence suggests a slightly greater susceptibility for women), and type of test (closed patch testing is decidedly more irritating than open). Season must also be considered although with closed patch testing the conditions are really equivalent throughout the year. In daily l-hour arm immersion tests, the skin is unquestionably more reactive in winter than summer, although the contrary impression is widely held. Justice 

### et al. (1961) showed this for surfactants, and we too have demonstrated greater winter reactivity for a variety of irritants. To obtain equivalent degrees of irritancy in summer it may be necessary to almost double the concentration.

In any case, irritancy data are valid only when the conditions are precisely specified. Even so, there remain trying technical problems which adversely affect accuracy and reproducibility. At present, it is unwise to suppose that the most exacting procedures yield values which have anything like the accuracy of clinical laboratory tests such as the serum cholesterol, glucose tolerance, or urinary ketosteroids. The most meticulously executed irritancy test yields values which are best regarded as relative rather than absolute. We must perforce rely on comparative evaluations. For example, in a two-way comparison ofproducts A and B, our currelat methods are adequate to decide whether A is significantly more irritating than B, less irritating than B, or indistinguishable. When a score of formulations are compared, those can usually be satisfactorily ranked in their order of irritancy. As a consequence, the products to be compared must be tested on the same subjects at the same time.

As a matter offact, comparative data are really quite reliable ifone makesjudicious
use of

### slandards ofreference. These are formulations or materials whose safety record is already known after widespread usage over a period of years. The usage history is a valuable item of information which can effectively be coordinated with the bioassay results in prognosticating toxicity. The first requirement is that the comparisons be between similar products: hand creams with hand creams, not with remedies for athletes foot; antiperspirants with antiperspirants, not with anti-dandruff proprietaries. The physical status should be the same, viz, liquids with liquids, etc. By the device of a reference standard, the artificial conditions ofbioassay may be related to actual usage.

To be feasible, bioassay must furnish accurate irritancy appraisals with small groups of test subjects. The objective is to obtain assessments in, say. 10 test subjects that would otherwise require perhaps 1 to 10,000 users to provide. Since with cosmetics we will usually be dealing with formulations of very low toxicity, this objective
can be attained only by tremendously magnifying or exaggerating the test exposures.

### 40 KLIGMAN



### The conditionsofbioassaybearnoresemblance to actualusage, anditmustbe stressed that theproportion ofsubjects irritatedbythetest system givesno directindicationof the probable incidence ofirritationintheuserpopulation. Since the details of our basic methods have alreadybeen published (Kligman and Wooding, 1967), it wilt sufficehere to outline our general approach. What I wish to emphasize is not the technical specifications, whichwillprobably be modifiedanyway as experience develops, butrather the strategies that canbe usedto compareirritativeness. We use small groups, usually 10 adults. Caucasians are preferable, but the experienced worker may utiIize more pigmented subjects. Theoreticallytwo irritancyparameters can be quantitated: (1) the proportion of subjects who react (frequency data) and (2) the degree ofreaction (intensity data). Appraising intensityis a highlysubjective and uncertain business with built-invagaries. The most important ofthese is the diversityofthe inflammatoryexpressions ofdifferentirritants so that uniformscoring becomes quite impossible. Many workers do not appear to be seriously troubled by the factthat above the erythemathreshold, irritants do not behave the same way. As the reactionbecomes stronger, avarietyofpicturesmaydevelop. Wehave simplybeen unable to force our observations into the conventional 4-point scale, so blithely describedin the literature: viz,+, erythema;++, deeper erythemaand swelling; ++% the foregoing plus vesicles or papules; ++++, the addition of bullae and crusting. Unfortunatelymany strong irritants never produce vesicles or bullae but simply kill the epidermis outright so that one gets necrosis, erosions, or ulcerationwithoutgoing through a blistering phase. Some irritants elicit pustules or induce suppuration. Among other possible manifestations are bloody, suppurative or serious oozing, various types of crusts, scaling, etc. Eight-point scales have been published in which the intensity oferythemaalone is graded over a 4-point range. Our main criticism of such scoring systems is notthat theycannotbe successfullyused by skilled observers, but rather that differentworkers are notlikelyto obtain comparable results. We have resolved this particular problem by ignoringit: we make no effortto grade intensity. Instead,we useprocedureswhich simplifyinterpretationby allowingthe observer only
one choice. Either uniformerythemaispresent or not; this is somewhat similarto the minimumerythemadose(MED)utilizedto estimatethe effectofultravioletradiation. The use of a quantal, all-or-none, decision increases reproducibility and greatly diminishes observer variability. It turns out thatthe accuracyofthetestincreases withthe potencyoftheirritant. It is comparativelysimple to appraise strong irritants which are capable ofcausing a reaction within24 hours in all subjects. The statisticutilized is the ID50: the concentration that will cause an erythema by occlusive patch testing in 50~o of the subjects. Various dilutions are employedover an appropriate range determinedby preliminary testing; an effort is made to obtain a good scatter. At the highest concentration 90-100 ~oshould react, and 0-10~oat the lowest. The percentage of subjects reactive to each concentration is plotted on log normal probability paper. A straight line is visuallyfitted and the ID50 is read offthe graph. Bymeans ofthe Litchfield-Wilcoxon method of probit analysis, one can establish the 95~ confidencelimits of the ID50, the slopefunctionandits confidence limits, and assessthe statisticalsignificance ofthe differences between irritants. One should note that substances which have identicalCOSMETIC IRRITANCY 

### 41

ID50 values are not necessarily ofequal irritancyunless their slope functions are also the same; the onewiththe steeper slope isthe strongerirritant.

For chemicals or formulations ofmoderate irritancy it is necessary to reapply the patch at the same site dailyfor 10 days, the end point being the number of days that willproduce an erythema. When this is reached no furtherpatches are applied to that site. The cumulative percentage of reactors per day is plotted against days on log normal probability paper as above (Fig. 3), The statistic provided in this case is the

99.99

99.9 99.S
99.5

99
98

95

90 SODIUM THIOGLYCOLLATE, 20% TIME- RESPONSE LINE

ITE4= 9.4days

80 B

Z 70

- 60 - • 0 IT50= 5.2 days 50
40 -- X

20 -- ITI6~ 2.9days



### ~ ~o0.2 0.I

0.01 1 2 3 4 5 6 7 8 910 20 30 40 50 607080 CUMULATIVE EXPOSURE TIME, d:,ys
Flo. 3. Log probitplot of the cumulative percentage of daily reactors in a 10-day continuous patch testwith sodium thioglycolate,The straightlinerepresents the best visualfit; the IT50 isthen directly read off.42 KLIGMAN

IT50, the number of days required tojust produce redness in 50~ ofthe subjects. The data are treated in the same way. With mildly irritating substances it may be necessary to extendthe daily exposuresto 20 daysin orderto exceeda 50~oreaction rate. Though
one might expect otherwise, it often happens that increasing the number of days fails to raise the reaction rate materially; for example if only 20~ react by the end of 10 days, another 10 days' exposure does not necessarily recruit additional reactors. With mild irritants it is almost characteristic for the cumulative response curve to level off within a week or so. 

### The Scotch Tape Stripping lrritancy Test The publication by Kligman and Wooding (1967), was limited to a description of the ID50 and IT50 procedures. Thesemethods have an important limitation: when the product or substance fails to induce reaction rates in excess of 50%, the IT50 or ID50 values cannot be reliably determined and the benefits of statistical analysis are lost. Regrettably, it turns out that these procedures are often inapplicable to cosmetics. A different strategy must be employed, to further magnify the exposure. The principle is similar to that used by Finkelstein 

### et aI. (1963) in animal testing. It entails mildly damaging the skin by chemical or physical means. The insult must be sufficient to disrupt the horny layer barrier and thus enhance the penetration rate. We tried various means of injury and finally decided on Scotch tape stripping, mainly because it can be precisely controlled, guarantees total removal of the barrier, and induces a uniform but mild inflammation. Our experienceis still too limited to lay down rigorous rules, but the results to date are promising enough tojustify a tentative description of the method. Since stripping itselfinduces some inflammation, one is obliged to forego quantal responses and to evaluate the intensity ofthe reaction. On stripped sites, even mild irritants will intensify inflammation. Obviously a Scotch tape stripping control, treated only with the vehicle, must be included. The sites are prepared by stripping to the glistening layer, using a uniform stripping technique. The material is applied continuously under occlusion once daily, generally for 5 days--less if great differences become apparent. We find that the intensitygenerallyreaches a maximum by 3-5 days and the skin then tends to recoverregardless offurther exposure. Intensity is arbitrarily scored on a 5-point scale by comparing the response to the control. Ten subjects are utilized, and the intensity readings are averaged for each day. Only experience can instruct the observer regarding rating of the reaction. Needless to say, this is a technical weakness of the test, and different observers are likely to score the reactions differently. With a suitable reference standard, however, a given observer can still reliably discriminate among weak irritants ofdiffering irritancy potential. The usefulness of the procedure may be illustrated by a comparison among a homologous series of straight-chain fatty acids. In 2 ~o concentration in mineral oil, these mimic many cosmetics in that their continuous occlusive application to normal skin for 10 days generally produces no reactors. Fatty acids were selected because it has been established that irritancy increases with chain length to a maximum at C12 and tends to fall thereafter (Kooyman and Snyder, 1942; Kellum, 1968). Figure 4 shows the results obtained by the stripping technique for even-numbered fatty acids from C8 to 

### C14, The comparative irritancies agree with the published results, attesting to the validity of the method. Means of handling these data statisticalIy have still to be worked out.COSMETIC IRRITANCY 

### 43

3 2% FATTY ACIDS

~- .... xcl4 A,---. __m C 12 o ........ o CIO ,i---- --A C 8
o--------~ C 6

..__.---- _ rj.



# ---j..j

~ I =,1 ;L I 2 ~ 4

Days
FiG. 4. Appraisal of the comparative irritancy offatty acids by the Scotch tape strippingmethod. Toxicityincreases with increasingchairt length to a maximum at Cz2. The separationis greatest at 2 days. 

### TechniquesofPatchTesting These bioassays uniformlyuse occlusivepatch tests, the details of which have been published elsewhere (Kligman and Wooding, 1967). Certain keypoints are worth reviewing and emphasizing. No test, ofcourse,isworth the troubleifitis slovenlyexecuted.It is thepeculiar pitfall ofthe patch test that one maycommit sins withoutrealizingthe transgression. The dangeristhatthepatchtest is

### beguiIinglysimple. Itsbasic mechanicsare easilylearned. One loads the patch and seals it to the skin with adhesive. However, it is the state of thepatch withinthe next 24 hours which is crucial. No warningis needed that the test isinvalidatedifthepatch comesofforisgrosslyloosened. Yetitissurprising how often the results are vitiated because ofseeminglyslightchanges in thephysical status ofthe patch. This occurs so frequently that it tends to be accepted as unavoidable, [ am referringto partial loss ofocclusion. Occlusionis absolutelyessentialin achievingreproducible results. The occlusivepatch is a device to provide continuous, uniform, intimate contact with the skin. This is not easy to achieve because the subjects are not immobilized and their normal physical activitystretches and deforms the skin. Movements during sleep can easily distort patches on the back. In a regional comparison Magnusson and Hersle (1965), thought the latter area was slightly more reactive because of pressure during sleep. The commonest type of loss of occlusion is wrinklingand foldingofthe patch. How many ofus would be willingto let an outside observer examine our next hundred subjects for integrity of the patch 24 hours after placement? Scrupulousness in applyingpatch tests so that theytruly remain occlusive for24hours is byfar the most decisive factorin achievingreliability. Our patches consist of 1-cm squares of Webril (Curity), a highly absorbent nonwoven fabric. The patches are loaded with a uniform quantity of the test solutions,
cream or ointmentfrom a tuberculin syringe.The patch is held tothe skinunder overlapping strips ofimpermeableplastictape (Blenderm, 3M). Finallythis arrangement is reinforced by overlapping longer strips of tape. A common error is to indulge the natural inclination to keep the entire dressing small. The strips of impermeable tape44 KLIGMAN

directIy over the cloth should be at 1east 2 inches long and the final outside tape at/east 3 incites long. These specifications may of course be individualized. Determination of erythema thresholds is more greatly influenced by lighting than
one might suppose. Fluorescent bulbs rich in blue light are especially handicapping. We use sunlight-mimicking light (Macbeth Corp., Newburgh, New York). Ifpossible, the summer months should be avoided. No dressing is secure in a stream of sweat. It has been reported that repeated patch testing leads to a gradual increase in skin irritability, resulting in abnormally high reaction rates even when completely different sites are used each time (van Haam and Mallette, 1955). Dermatologists generally believe that the skin becomes hyperirritable in the presence of a widespread dermatitis, but except for Bj/3rnberg's (1968) suggestiveresults, I know ofno telling proof. Indeed, I have repeatedly tried to confirm this nonspecific increase in irritability without success. For example, if a severe dermatitis on both forearms is induced by daily immersions in irritating chemicals the ID50 values on the back at the very height of inflammation were not found to be different from the preexposure figure. In addition we have repeatedly used the same subjects for testing strong irritants over a period of several months, without noticing any change in skin susceptibility. I am ofthe opinion, therefore, that when one has assembled a cooperative panel of test subjects, these can be used indefinitely. It is probably a sound precaution to leave several weeks between tests and to avoid sites previouslyirritated.

REFERENCES BJrRNB~RO, A. (1968). Skin. Reaetio1~s to Primary Irritants in Patients with Hand Eczema. Doctoral thesis. Oscar Isacsons Tryckeri, Goteburg, Sweden. CA.LNAN,"D.C., and SRUSTeR,S. (1963). Reactions to ammoniumpersulfate.Arch. Dermatol. 88, 12. FnqKLESTEIN,P., LADEN,K., and 

### MIECHOWSKI, W._(1963).New methods for evaluating cosmetic irritancy, a

v.h~vest. DermatoI. 40, 11. JUSTICe,J. D., Te,

hVERS,J. J., and VrNSON,L. J. (1961). The correlation between animal and human tests in assessingproduct mildness. Proc. Sci. Sect. Toilet Goods Assoc. 35, 12. 

### KELLUM,R. E. (1968). Ache vulgaris. Relative irritancy of four fatty acids from C12 to CI6. Arch DermatoL 97, 722. Kt.x~MA~q,A. M., and WOODING, W. M. (1967). A method for the measurement and evaluation of irritants on human skin. J. hwest. Dermatol. 49, 78. KOOYMAN,D. J., and StqvDta, F. A. (1942). Tests for the mildness ofsoaps. Arch. Dermatol. 46, 846. 

### MAGNUSSON,

B., and HE~SLE,K. (1965). Patch test methods. II. Regional variations ofpatch test responses. Acta Dermato-Venereol. 45, 257. S~ELANSKI, H. A. (1951). Experiences with and considerations of the human patch test method. 3. Soc. Cosmet. Chem. 11, 324. vc3NHAHN, E., and MALUETrE, F. S. (1955). Skin testing of industrial compounds. Arch. b~cl.Hyg. 16, 324. WAgSHAW,T. S., and HERr~MArqN,F. (1951). Studies of skin reactions to propylene glycol,
o r. Invest. Dematol. 19, 423.