Guidance for Reviewers

Potency Limits for Standardized Dust

Mite and Grass Allergen Vaccines:

 A Revised Protocol

 Additional copies are available from:

 Office of Communication, Training and Manufacturers Assistance (HFM-40)

 1401 Rockville Pike, Rockville, MD 20852-1448 

 (Tel) 800-835-4709

 or 301-827-1800 

 (Internet) http://www.fda.gov/cber/guidelines.htm

U.S. Department of Health and Human Services

Food and Drug Administration

Center for Biologics Evaluation and Research

November 2000
i

Table of Contents

[Note: page numbers may vary for documents distributed electronically.]

I. INTRODUCTION..................................................................................................................... 1

II. DIAGNOSTIC EQUIVALENCE............................................................................................ 2

III. LITERATURE REVIEWED ................................................................................................. 2

IV. THERAPEUTIC EQUIVALENCE........................................................................................ 2

V. SAFETY EQUIVALENCE....................................................................................................... 3

A. Linear Regression........................................................................................................ 3

B. Logistic Regression...................................................................................................... 3

C. Safety Equivalence: Summary..................................................................................... 4

VI. LOT-TO-LOT VARIATION IN ALLERGEN VACCINE POTENCY............................... 4

A. Sample Variance............................................................................................................ 5

B. Potency Range............................................................................................................... 5

VII. CURRENT CBER RELEASE LIMITS .............................................................................. 6

VIII. BROADENED RELEASE LIMITS................................................................................... 7

IX. REFERENCES ....................................................................................................................... 8

TABLE 1....................................................................................................................................... 10

TABLE 2....................................................................................................................................... 11

TABLE 3....................................................................................................................................... 12

TABLE 4....................................................................................................................................... 13
1

GUIDANCE FOR REVIEWERS

Potency Limits for Standardized Dust Mite and

Grass Allergen Vaccines: A Revised Protocol

This guidance document represents the agency’s current thinking with regard to potency limits for standardized

dust mite and grass allergen vaccines. It does not create or confer any rights, privileges, or benefits on or for any

person and does not operate to bind FDA or the public. An alternative approach may be used if such approach

satisfies the requirements of the applicable statutes and regulations.

I. INTRODUCTION

The release limits for standardized dust mite and grass allergen vaccines are based on the performance

characteristics of the competition Enzyme-Linked Immunosorbent Assay (ELISA) for the determination

of the relative potency (RP) of these products. Using this assay, Center for Biologics Evaluation and

Research (CBER) has required (21 CFR 680.3(e)) that all lots of these products be shown to be

equivalent to the reference standard, with 95% confidence. Thus, the limits have been set at 0.70-1.43

for three determinations; when six determinations are made, the limits are 0.78-1.29.

These release limits will remain unchanged. However, CBER is establishing broader RP limits of 0.5 to

2.0 for its evaluation of standardized dust mite and grass allergen vaccines submitted to CBER for lot

release. CBER lot release evaluations will be carried out with three replicates, using the competition

ELISA recently re-validated by CBER. The manufacturers will be expected to maintain their current lot

release limits, as detailed in their approved license applications. While these internal limits will continue

to impose equivalence to reference at 95% confidence with either 3 or 6 replicates, CBER will only fail

lots that, in its laboratory, fall outside the 0.5 to 2.0 limits.

In establishing broader release limits for CBER’s testing of submitted allergen vaccines, CBER

recognizes the predictable uncertainty associated with the submission of a product with a RP at or near

the previously acceptable limits. The new CBER limits of 0.5 – 2.0 are statistically equivalent to

applying a 95% confidence limit on the previous release limits for n = 3.

The potency limits for standardized allergen vaccines (Ref. 1, 2) should be based on acceptable ranges

established in clinical studies. Three criteria appear to be important. The first, therapeutic

equivalence, addresses the efficacy of allergen vaccines for immunotherapy. Thus, a RP range will have

the property of therapeutic equivalence if, for the allergen vaccine in question, lots with relative

potencies anywhere in that range have an equal likelihood of effecting clinical improvement in an

immunotherapy trial. Likewise, diagnostic equivalence addresses the efficacy of allergen vaccines for

in vivo diagnostics. Finally, safety equivalence reflects the likelihood of the safe administration of the

vaccine for either diagnostic or therapeutic indications. The limits acceptable to CBER should fall within

the narrowest of the equivalence ranges established by these criteria.
2

These clinical limits are discussed in sections II. through V. of this document. In addition, the variability

of the potency of allergen vaccines can be considered in setting safe release limits; this is discussed in

section VI. Finally, the current and broadened release limits are discussed in sections VII. and VIII. 

This guidance document finalizes the draft guidance document entitled "Guidance for Reviewers:

Potency Limits for Standardized Dust Mite and Grass Allergen Vaccines: A Revised Protocol" that was

announced in the Federal Register on February 15, 2000 (65FR 7557).

II. DIAGNOSTIC EQUIVALENCE

The range of diagnostic equivalence is broad. The overwhelming majority of allergists use a qualitative

grading system based upon the wheal size after percutaneous or intracutaneous skin testing (Ref. 3). In

one study, the mean slope of the skin test titration curve was only 2.7 mm per 3-fold dose dilution for

wheals. When standardized 10,000 and 100,000 BAU/mL grass extracts were compared, wheal size

varied only minimally, but achieved statistical significance (P. Turkeltaub, unpublished data). For

allergists who use erythema skin testing, the minimum detectable variability is only 3-4 fold (Ref. 4, 5) .

III. LITERATURE REVIEWED

Seven studies were chosen for analysis of therapeutic and safety equivalence, six of Amb a 1, one of

Der p 1 (Ref. 6-12) (Table 1). Only three of the studies utilized allergen extracts that were rigorously

standardized (Ref. 8-10) and three lacked any appropriate controls (Ref. 7-9). Nonetheless, each

study provided some dose-response data on the efficacy and/or safety of allergen immunotherapy,

although one (Ref. 9) was selected because of its observations on the physiologic effects of low dose

immunotherapy. Six were analyzed for determination of therapeutic equivalence (Ref. 7-12) and three

for determination of safety equivalence (Ref. 6,10, 12).

IV. THERAPEUTIC EQUIVALENCE

The range of equivalent therapeutic doses of allergen vaccines appears to be broad, and few

investigators have performed detailed dose response evaluations in immunotherapy. Common allergy

practice is to target the “maximum tolerated dose” as the therapeutic objective; on the other hand,

controlled studies have suggested that maintenance dosing over a four-fold range of antigen is

therapeutic (Ref. 1, 2, 13). Dose responses, when they have been performed, have been inexact. In

each of the six studies chosen for this analysis, the investigators found that well-characterized allergen

vaccines containing Amb a 1 or Der p 1 had a therapeutic effect over a broad dose range (Table 1). In

three of the studies, symptom score improvements were examined after patients attained different

allergen doses in their treatment. One study showed equivalence over a two-fold range (Ref. 8); another

over a 14-fold range (Ref. 10); and the third over a 30-fold range (Ref. 12). Similarly, those studies

that measured physiologic changes associated with allergen immunotherapy suggested equivalent

responses over 12-fold (Ref. 11) and 1000-fold ranges (Ref. 7). The study by Hedlin et al., (Ref. 9) is

notable in that multiple physiologic and immunologic changes associated with successful allergen

immunotherapy appeared after achieving a dose of Amb a 1 of only 0.11 μg (cumulative dose 0.22 μg),

a dose at least 10-fold less than what is normally associated with successful allergen immunotherapy.
3

V. SAFETY EQUIVALENCE

On the other hand, the range of safety equivalence of allergen vaccines may be narrower. In the

recent WHO position paper on allergen immunotherapy, panel members distinguished between the

optimal dose and the maximum tolerated dose, and noted that the range between these may be highly

variable (Ref. 1, 2).

Among the three selected studies that addressed the occurrence of systemic adverse reaction rates at

different doses of allergen (Ref. 6, 10, 12), two included subsets of patients that warranted separate

analysis. Van Metre et al., (Ref. 6) placed fifteen study subjects on standard, weekly immunotherapy,

and eighteen subjects on a cluster regimen. Turkeltaub et al., (Ref. 10) described systemic reaction rates

at various doses, as well as the rates of reactions severe enough to require treatment with subcutaneous

injection of epinephrine. In our analysis of these two studies, these subsets were considered separately,

even though there was some overlap of the patient groups described by Turkeltaub et al.

Further, some of the data were analyzed by the investigators on a per injection basis, and others on a

per patient basis. Since these data cannot be combined or compared, they have been treated

separately.

A. Linear Regression

For this analysis the adverse reaction rate is fitted against log dose. The fitted slopes and their

standard errors are listed in Table 2. Thus, as a first estimate, one would expect a 4-11%

increase in systemic reaction rates associated with each injection of an extract containing a tenfold greater extract potency than the previous injection; on a per patient basis, the rise in

systemic reaction rate would be 16-25%.

A more precise estimate is obtained by pooling the preceding slopes with weighting in inverse

proportion to their standard errors. This yields an increase in the systemic reaction rate of 5.9%

(per injection) or 19.6% (per patient) for a ten-fold increase in allergen dose.

B. Logistic Regression

To account for the fact that probabilities are bounded by 0 and 1 and tend to saturate at the

extremes, fitting was also carried out with the logistic model

m d b

p

p

i

i

i

= + 















−

ln( )

1

ln

where piis the probability of a reaction, and diis the dose. The interpretation of the fitting

parameters necessarily differs from the prior analyses, in that incremental changes in reaction

rates are not constant in all dosage ranges. Thus, a reasonable mean dose must be specified,

and the increase in dose that would produce a given increase in systemic reaction rate

calculated.
4

Defining d0 as the geometric mean of the high and low doses, p0, the probability of an adverse

reaction at dose d0, can be calculated directly from the equation after m and b have been

determined. We then define the dose at which p0 would be expected to increase by 0.05 as

d0.05 . This can be evaluated from the logistic equation:

m d b

p

p

= + 















−

+

ln( )

0.95

0.05

ln 0.05

0

0

The results of the fitting and subsequent determination of the ratio d0.05 d0 are listed in Table

3. This analysis shows increasing the dosage between 2.4 and 5-fold (an average of 4-fold)

leads to a 5% increase in the adverse reaction rate for the per injection data; for the per patient

data, 1.5 to 2-fold (an average of 1.7) dose increases lead to 5% adverse reaction increases.

C. Safety Equivalence: Summary

Unfortunately, little else has appeared in publication that can shed light on the question of safety

equivalence. While the data in Table 1 indicate that increases in RP typically lead to increases in

adverse systemic reactions to immunotherapy, the range is quite varied and the study designs

are suboptimal. Clearly, studies with better design (placebo-controlled, double blind with welldefined, highly sensitive subjects and well-characterized products with FDA acceptable

standard of potency) would be preferable and are needed.

It should be noted that only safety data within therapeutically equivalent dose ranges were

analyzed. Thus, while all of Haugaard’s (Ref. 12) data were included for analysis, the lowest

data points in Turkeltaub (Ref. 10; 0.003 μg) were excluded.

It is not possible, based on these studies alone, to assign a definite value to the increased risk of

systemic reaction associated with increased potency. A conservative estimate of an acceptable

increase in potency may be attempted, if we assume that the increase in the adverse reaction

rate upon an increase in dosage should be less than 5%, and that the final reaction rate should

not be much greater than 5-10%. Reaction rates in this range may be an unavoidable feature of

curative immunotherapy with native antigens. At best, this analysis shows that the clinical data

available suggest that a four-fold variation in allergen vaccine potency yields an acceptably small

increase in systemic reaction rates.

VI. LOT-TO-LOT VARIATION IN ALLERGEN VACCINE POTENCY

The aggregate consistency of manufactured lots might also be taken into account when establishing

testing limits. For example, if typical lot-to-lot consistency were very high and well within clinical limits,

then testing protocols could be adjusted to eliminate outliers while rarely failing lots whose RP is close to

unity. On the other extreme, if the distribution of lots were broad, then stastical equivalence to the

reference would be appropriate.
5

A. Sample Variance

The observed variability of a group of lots of an allergen vaccine is a function of the sample

variability and the variability of the assay. Defining the densities of the sample, assay and

observed distributions f (x)

s

, f (x)

a

, and f (x)

obs , respectively, the relation between them is

(Ref. 14):

∫

∞

−∞

f x = f t f x − t dx obs s a( ) ( ) ( )

Assuming that the sample and assay densities are normal with variances 2 ssand 2 sa,

respectively, f (x)

obs is normal with variance

2 2 2 sobs = ss +sa

An estimate of the variability of lots submitted to CBER can be obtained from the failure rate of

the submitted products and the intrinsic variability of the competition ELISA that was used to

test the products. Between 1995 and 1997, ten different manufacturers submitted 412 lots of

grass pollen extract in support of licensure. Likewise, between 1995 and 1999, eleven

manufacturers submitted 91 lots of licensed dust mite extracts for approval. The failure rates of

these lots were analyzed to determine lot variability.

For the grasses, 51 of 412, or 12.4%, failed lot release specifications. Of these, 29 were

above the upper limits, and 22 fell below the lower limits. Assuming that the submitted lots are

normally distributed about the reference (in x = log RP), half would have failed high implying

that 93.8% were below the upper limit, x'= log1.53 = 0.1847 :

∫−∞

=

'

( ) 0.938

x

f obs x dx

The standard deviation of obs f may be obtained by transforming to the standard normal, in

which case, obs z'= x'/s . From a table of the cumulative normal distribution, z'=1.54, or

sobs = 0.120. The standard deviation of the assay with three replicates, sa is

0.1375/ 3 = 0.0794. It then follows that ss = 0.090 .

For standardized dust mite allergen vaccines, 6 of 91 failed (3 high and 3 low), the preceding

analysis yields z'=1.84 and σs = 0.061.

B. Potency Range

The change in RP when switching bottles can be estimated from the absolute difference of two

samples picked at random from a distribution. This is a special case of determining the range,

R, which is defined by the difference between the highest and lowest of n samples taken from a
6

distribution with density f(x) (Ref. 14). For n=2, the density, f (r) R, is given by









<

+ >

=

∫

∞

−∞

0 0

2 ( ) ( ) 0

( )

r

f x f r x dx r

f r R

The average range is then calculated from

∫

∞

=

0

r r f (r)dr R

It is also useful to denote the quantity r' such that

( ) 0.95

'

0

= ∫

r

f Rr dr

That is, 95% of the values of the range are less than r' .

For a normal (or Gaussian) density with variance 2 s , the density, average, and 95% maximum

for the range are as follows:

( ) ( )

2 2

1

f R(r) = ps exp − r / 4s

−

r = (2/p)s ≈ 0.8s

and r'= 2 ×1.96s ≈ 2.8s

From the values of σ calculated above, r = 0.0718 and r'= 0.249 for the grass extracts.

These ratios are in log RP. Thus, the anticipated mean change in RP upon switching bottles is

(100.0718 - 1), or 18%. Likewise, 95% of the potency shifts will be less than (100.249 - 1), or

80%.

For the mites, r = 0.049, and r' = 0.171, implying that the mean change in RP is 12%, and

that potency shifts will be less than 48% in 95% of cases.

VII. CURRENT CBER RELEASE LIMITS

Given the uncertainty of the true clinical limits for the accuracy of allergen vaccine content, CBER has, in

the past, utilized the accuracy of the test used to measure RP as the de facto limits for vaccine approval.

 Thus, the 95% Confidence Interval (CI) limits for the Radioallergosorbent Test (RAST) inhibition assay

were 0.46 to 2.12. These limits were acceptable since they were equivalent to the erythema diagnostic

equivalence data. The competition ELISA is more accurate and, using this assay, CBER narrowed the
7

95% CI limits to 0.70-1.43 for three determinations; when six determinations are made, the limits are

narrowed further to 0.78-1.29. The strength of CBER’s current approach is that it decreases the

likelihood that a mite or grass allergen vaccine with a RP substantially different from 1.0 will be released

in the US market.

VIII. BROADENED RELEASE LIMITS

The internal release limits that establish equivalence to the reference at 95% confidence will remain

unchanged. We are establishing RP limits of 0.5 to 2.0 for standardized dust mite and grass allergen

vaccines submitted to CBER for lot release. CBER lot release evaluations will be carried out with three

replicates, using the competitive ELISA recently validated by LIB. The manufacturers will be

expected to maintain their current lot release limits. These internal limits now impose equivalence

to reference at 95% confidence with either 3 or 6 replicates, depending on the manufacturer. CBER will

consider as acceptable lots that fall within the 0.5 to 2.0 limits. This represents an expansion of the RP

limits that CBER would find acceptable.

In initiating this change, we also expand the RP limits for the shelf life of the product, provided the initial

release data fall within the internal lot release specifications. Current shelf life limits are 0.568-1.759, as

follows from a Bonferroni correction (Ref. 15, 16). Hence, the expansion to 0.5 to 2.0 allows for a

small degree of decay over the shelf life of the standardized allergen vaccines currently licensed. 

Current data suggest that these glycerinated vaccines are stable; if further data confirm this stability over

the current three-year dating period, the acceptable manufacturers’ internal limits may be broadened.

The likelihood of lot failure by CBER depends on the manufacturer’s internal specifications (the limits

and the number of internal replicates). A product subjected to 6 replicate evaluations by the

manufacturer, with a result falling between 0.776 and 1.288, has no greater than a 2.5% chance of

failure. On the other hand, a product subject to only three replicates has a 9.8% chance of failure (Table

4).

While the preponderance of studies suggests that a ten-fold range of RP would be acceptable from a

therapeutic point of view, we believe that the four-fold range is preferable because of safety concerns. 

Furthermore, future studies comparing the efficacy of allergen vaccines will be difficult to interpret if a

ten-fold range of potency becomes the accepted norm. Finally, the four-fold range is well within the

established capabilities of the allergen manufacturers using current techniques and standards.

The 0.5-2.0 limits are the same as those proposed by the European Union (EU) and, thus, nominally

achieve harmonization. However, there are important differences. The procedure established here

includes tighter internal limits on the manufacturer and follows with a confirmatory test by CBER. This

greatly reduces the probability that the true RP of any product falls outside the 0.5-2.0 range, even if it

were not tested by CBER. The EU document, in contrast, does not stipulate either test methodology

or confidence level. Hence, according to the EU standard, a lot tested by the manufacturer as 0.51 is

acceptable as long as it passes subsequent testing by the appropriate regulatory agency. This would not

be the case using the limits described in this guidance document.

In establishing broader release limits for CBER’s testing of submitted allergen vaccines, we recognize
8

the predictable uncertainty associated with the submission of a product with a RP at or near the

previously acceptable limits. The new CBER limits of 0.5 – 2.0 are statistically equivalent to applying a

95% confidence limit on the previous release limits for n = 3. Thus, the 95% confidence lower limit of

an allergen vaccine with a RP of 0.699 is 10log 0.699-1.96σ/√n, or 0.488. Likewise, the 95% confidence

upper limit of an allergen vaccine with a RP of 1.431 is 10log 1.431+1.96σ/√n, or 2.047.

The advantages of adopting these broadened release limits are:

1. We establish a system in which the limits are based upon the human clinical response to allergen

vaccines.

2. We differentiate between limits based on clinical data and those based on the accuracy of the in

vitro assays used to measure RP. This distinction will be especially important as our in vitro methods

become more accurate in the years to come.

3. The 0.5 to 2.0 limits are within the previously acceptable limits based on the RAST inhibition assay.

These broader limits were not associated with any increased safety concerns.

4. Manufacturers that perform six replicate evaluations are extremely unlikely to experience lot

rejection by CBER.

IX. REFERENCES

1. Allergen immunotherapy: therapeutic vaccines for allergic diseases. Geneva: January 27-29 1997. 

Allergy 1998; 53:1-42.

2. Bousquet J, Lockey R, Malling HJ. Allergen immunotherapy: therapeutic vaccines for allergic

diseases. A WHO position paper. J Allergy Clin Immunol 1998; 102:558-562.

3. Bernstein IL, Storms WW. Practice parameters for allergy diagnostic testing. Joint Task Force on

Practice Parameters for the Diagnosis and Treatment of Asthma. The American Academy of

Allergy, Asthma and Immunology and the American College of Allergy, Asthma and Immunology.

Ann Allergy Asthma Immunol 1995; 75:543-625.

4. Turkeltaub PC, Rastogi SC, Baer H, Anderson MC, Norman PS. A standardized quantitative

skin-test assay of allergen potency and stability: studies on the allergen dose-response curve and

effect of wheal, erythema, and patient selection on assay results. J Allergy Clin Immunol 1982; 

70:343-352.

5. Turkeltaub PC. Biological standardization. Arb Paul Ehrlich Inst Bundesamt Sera Impfstoffe

Frankf A M 1997; 145-156.

6. Van Metre TE, Jr., Adkinson NF, Jr., Amodio FJ, Kagey-Sobotka A, Lichtenstein LM, Mardiney

MR, Jr., et al. A comparison of immunotherapy schedules for injection treatment of ragweed

pollen hay fever. J Allergy Clin Immunol 1982; 69:181-193.

7. Creticos PS, Van Metre TE, Mardiney MR, Rosenberg GL, Norman PS, Adkinson NF, Jr. Dose
9

response of IgE and IgG antibodies during ragweed immunotherapy. J Allergy Clin Immunol 1984;

 73:94-104.

8. Creticos PS, Marsh DG, Proud D, Kagey-Sobotka A, Adkinson NF, Jr., Friedhoff L, et al. 

Responses to ragweed-pollen nasal challenge before and after immunotherapy. J Allergy Clin

Immunol 1989; 84:197-205.

9. Hedlin G, Silber G, Schieken L, Naclerio R, Proud D, Eggleston P, et al. Attenuation of allergen

sensitivity early in the course of ragweed immunotherapy. J Allergy Clin Immunol 1989; 84:390-

399.

10. Turkeltaub PC, Campbell G, Mosimann JE. Comparative safety and efficacy of short ragweed

extracts differing in potency and composition in the treatment of fall hay fever. Use of allergenically

bioequivalent doses by parallel line bioassay to evaluate comparative safety and efficacy. Allergy

1990; 45:528-546.

11. Furin MJ, Norman PS, Creticos PS, Proud D, Kagey-Sobotka A, Lichtenstein LM, et al. 

Immunotherapy decreases antigen-induced eosinophil cell migration into the nasal cavity. J Allergy

Clin Immunol 1991; 88:27-32.

12. Haugaard L, Dahl R, Jacobsen L. A controlled dose-response study of immunotherapy with

standardized, partially purified extract of house dust mite: clinical efficacy and side effects. J Allergy

Clin Immunol 1993; 91:709-722

13. Weber RW. Immunotherapy with allergens. JAMA 1997; 278:1881-1887.

14. DeGroot MH. Probability and statistics. London: Addison-Wesley, 1975.

15. Pagano M, Gauvreau K. Principles of biostatistics. Belmont, California: Duxbury Press, 1992.

16. Draft Guidance for Industry: Testing limits in stability protocols for standardized grass pollen

extracts. August 1997; Food and Drug Administration, Center for Biologics Evaluation and

Research.
10

TABLE 1

Summary of cited studies

Allergen Dose response

endpoint(s)

Number of

patients in active

group

Dose range Observations Reference Notes

Amb a 1 Systemic reactions 33 Up to 18.7 μg 7/15 patients undergoing the weekly

regimen, and 10/18 patients

undergoing the cluster regimen,

experienced systemic reactions at

doses ranging from 0.13 to 13.1 μg.

6

Van Metre, et al. 1982

Commercial lyophilized product,

compared with purified reference

allergens by RID. Placebo control.

Amb a 1 Antibody responses 51 Up to 93.5 μg Threshold doses for antibody

responses varied 1000-fold.

7

Creticos et al.

1984

Commercial aqueous extract.

Standardization uncertain. No

placebo or untreated control.

Amb a 1 Symptom scores and

nasal challenge

11 0.6, 6 and 12 μg 0.6 subtherapeutic; 6 and 12

equivalent and effective.

8

Creticos et al.

1989

Aqueous product prepared by

investigators from ragweed pollen,

and compared with CBER reference

standard by RID and crossed

immuno-electrophoresis. No placebo

or untreated control.

Amb a 1 Nasal challenge and

antibody responses

40 Up to 0.11 μg Measurable decreases in Amb a 1-

induced nasal histamine and TAME

release; decrease in skin test

reactivity; and increase in ragweedspecific IgE after a cumulative Amb a

1 dose of only 0.22 μg.

9

Hedlin et al.

1989

Commercial aqueous extract, defined

Amb a 1 content. No placebo or

untreated controls.

Amb a 1 Symptom scores and

systemic reactions

129 0.003, 0.3, 1.8, 2.25

and 4.2 μg

0.003 dose ineffective; all other doses

effective. Systemic reaction rate

(reactions/ injection) using standard

protocol: 2.1% at 0.8 μg and 5.6% at

4.2 μg. Rush protocol: 2.3% at 0.003

μg, 2.8% at 0.3 μg, 22% at 2.7 μg,

11% at 4.3 μg. Percent of patients

requiring epinephrine: 7.5% when the

maximum dose was 0.3 μg, 15% at

0.82 μg, 23% at 2.7 μg, 30% at 4.2

μg, and 25% at 4.3 μg.

10

Turkeltaub et al.

1990

Aqueous products analyzed by RID,

RAST inhibition and parallel-line

bioassay, and standardized by

comparison with CBER reference

standard. Untreated control, no

placebo control.

Amb a 1 Seasonal and postchallenge nasal

eosinphilia

89 2 and 24 μg High and low doses effective in the

challenge phase of study. In the

seasonal phase, only the higher dose

was effective.

11

Furin et al.

1991

Source and standardization of

ragweed extract uncertain. Untreated

control, no placebo control.

Der p 1 Symptom scores and

systemic reactions

81 0.7, 7 and 21 μg All three doses therapeutically

equivalent. Systemic reaction rate

(reactions/injection) 0.56% at 0.7 μg,

3.30% at 7 μg, and 7.10% at 21 μg.

12

Haugaard et al.

1993

Commercial aqueous (skin testing)

and alum adsorbed (IT) extracts. 

Compared to an internal standard by

RAST inhibition,

immunoelectrophoresis, and bioassay

(HEP method). Untreated control,

no placebo control.
11

TABLE 2

Slopes and their standard errors (SE) from linear fits of adverse reaction rates vs. log dose. Unless

otherwise noted in the text, all data from each report were analyzed. The final 2 columns list weighted

averages (over all sources for each study design) for the % increase in adverse reaction rates expected

for ten-fold and four-fold increases in dose, respectively. 

Study design Source Slope SE % increase in

adverse reaction

(ten-fold dose)

% increase in

adverse reaction

(four-fold dose)

Haugaard et al.

1993 (12)

4.2 1.3

Haugaard et al.

1993 (12)

(maintenance)

Per injection 9.1 3.8

Turkeltaub et al.

1990 (10)

11.1 10.9

5.9 3.5

Van Metre et al.

1982 (6)

(weekly)

25.3 2.4

Van Metre et al.

1982 (6)

(cluster)

Per patient 16.4 2.0

Turkeltaub et al.

1990 (10)

(epinephrine)

17.2 2.2

19.6 11.8
12

TABLE 3

Summary of logistic regression. Fitting parameters, m and b are defined by Eq. (1); d0 is the geometric

mean of the highest and lowest doses used in each study; and p0 is the probability of an adverse

reaction at d0calculated from Eq. (1). As discussed in the text and defined by Eq. (7), d0.05 is the

estimated dose in which the probability of an adverse reaction increases by 0.05 over p0. Units of m,

b and d0 are omitted for clarity.

Study design Source m b d0 p0 d0.05 d0

Haugaard et al.

1993 (12)

0.77 -4.98 4.69 0.02 4.6

Haugaard et al.

1993 (12)

(maintenance)

Per injection 1.12 -5.11 4.69 0.03 2.4

Turkeltaub et al.

1990 (10)

0.29 -2.33 1.52 0.10 5.0

Van Metre et al.

1982 (6)

(weekly)

0.67 -2.11 4.25 0.24 1.5

Van Metre et al.

1982 (6)

(cluster)

0.29 -0.49 4.25 0.48 2.0

Per patient

Turkeltaub et al.

1990 (10)

(epinephrine)

0.56 -1.76 1.64 0.19 1.7
13

TABLE 4

Probability that CBER will pass or fail an allergen vaccine with a submitted RP (RP) of 0.5 to 2.0. 

Note that the proposed standard will require manufacturers to continue submission of vaccines within

the 95% CI of RP = 1, indicated in bold typeface, for Nmanu = 3, or for Nmanu = 6.

N(manu) = 3 N(manu) = 6

RP P(fail) low P(fail) high P(pass) RP P(fail) low P(fail)

high

P(pass)

0.5 0.500 0.000 0.500 0.5 0.500 0.000 0.500

0.6 0.240 0.000 0.760 0.6 0.208 0.000 0.792

0.699 0.098 0.000 0.902 0.7 0.066 0.000 0.934

0.7 0.097 0.000 0.903 0.776 0.025 0.000 0.975

0.8 0.035 0.000 0.965 0.8 0.018 0.000 0.982

0.9 0.011 0.001 0.988 0.9 0.004 0.000 0.995

1 0.004 0.004 0.993 1 0.001 0.001 0.998

1.1 0.001 0.010 0.988 1.1 0.000 0.004 0.996

1.2 0.000 0.024 0.976 1.2 0.000 0.011 0.989

1.3 0.000 0.048 0.952 1.288 0.000 0.025 0.975

1.4 0.000 0.084 0.916 1.3 0.000 0.027 0.973

1.431 0.000 0.098 0.902 1.4 0.000 0.056 0.944

1.5 0.000 0.133 0.867 1.5 0.000 0.099 0.901

1.6 0.000 0.194 0.806 1.6 0.000 0.159 0.841

1.7 0.000 0.265 0.735 1.7 0.000 0.234 0.766

1.8 0.000 0.342 0.658 1.8 0.000 0.319 0.681

1.9 0.000 0.421 0.579 1.9 0.000 0.409 0.591

2 0.000 0.500 0.500 2 0.000 0.500 0.500

P(pass) = f (x)dx

log .

log .

0 5

2 0

∫

where x is the log of the RP calculated by the manufacturer (with N manureplicates) and subsequently by

CBER (with 3 replicates). f(x) is a normal distribution in log RP with variance,

s s s

2 2 2

2 2

01375

3

01375

= +

= +

CBER manu

N manu

( . ) ( . )

and 0.1375 is the standard deviation in log RP of the current CBER ELISA.