Measles-Virus Neutralizing Antibodies in Intravenous Immunoglobulins

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1 MAJOR ARTICLE Measles-Virus Neutralizing Antibodies in Intravenous Immunoglobulins Susette Audet, 1 Maria Luisa Virata-Theimer, 2 Judy A. Beeler, 1 Dorothy E. Scott, 2 Douglas J. Frazier, 2 Malgorzata G. Mikolajczyk, 2 Nancy Eller, 2 Feng-ming Chen, 2 and Mei-ying W. Yu 2 Divisions of 1 Viral Products and 2 Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland Measles infection induces lifelong immunity; however, wild-type infection stimulates higher levels of measlesvirus neutralizing antibodies (mnabs) than does vaccination. Because the proportion of the donor population with vaccine-induced measles immunity is increasing, this study was conducted to determine whether this shift in demographic characteristics affects mnab levels in contemporary lots of Immune Globulin Intravenous (Human) (IGIV). When 166 lots of 7 IGIV products manufactured between 1998 and 2003 were assayed by plaque-reduction neutralization test, there was a progressive decrease in geometric mean titers in lots manufactured between 1999 and IGIV products manufactured from recovered plasma had significantly higher titers than did those manufactured from Source Plasma, which could reflect a change in donor demographic characteristics, because Source Plasma donors tend to be much younger. A reduction in mnabs also correlated with the loss of either IgG1 and IgG3, possibly because of certain manufacturing procedures, or bivalent antibodies (i.e., intact IgG and F(ab ) 2 ), because of fragmentation. Immune Globulin Intravenous (Human) (IGIV) became available in 1981, and 9 products are currently licensed in the United States. These products are the mainstay for treating nearly 50,000 patients who have primary immunodeficiency disorders (PIDD) [1] and who rely on immunoglobulin prophylaxis to prevent common infectious diseases [2, 3]. Many factors may have an impact on the quality and quantity of antibodies in immunoglobulin products. Each lot is prepared from plasma pools derived from as many as 60,000 donors, to ensure that a large repertoire of antibodies is present in the final preparation [4]. Also, to be approved by the Food and Drug Administration (FDA), both Immune Globulin (Human) (IG) products and IGIV products must meet minimum potency requirements (which are provided in FDA reg- Received 28 December 2005; accepted 21 April 2006; electronically published 16 August Potential conflicts of interest: none reported. The views presented in this article do not necessarily reflect those of the Food and Drug Administration. Reprints or correspondence: Dr. Mei-ying W. Yu, FDA/CBER, 1401 Rockville Pike (HFM-345), Rockville, MD (mei-ying.yu@fda.hhs.gov). The Journal of Infectious Diseases 2006; 194:781 9 This article is in the public domain, and no copyright is claimed /2006/ $15.00 ulation 21 CFR ) for antibodies against measles, diphtheria, and poliovirus type 1, 2, or 3. Fluctuations in the measles-virus neutralizing antibody (mnab) potency of immunoglobulin products may reflect changes in the epidemiology of measles in the United States. Recent seroprevalence surveys suggest that 93% of the US population has measles antibodies [5]; this high level of herd immunity limits measles circulation in the United States, and, as a result,!100 measles cases/year were identified between 2001 and 2003, with no evidence of endemic measles transmission since 1997 [6]. Individuals who have experienced wild-type measles infection tend to have higher mnab titers than do those who acquire immunity by vaccination [7, 8]. Although there is a high prevalence of measles antibodies in the US population, important differences in the origins of this immunity may eventually affect the potency of immunoglobulin products. The aim of the present study was to survey mnabs in IGIV products, as a way to examine trends over successive years of manufacture and to see whether factors related to manufacture might influence potency. Finally, peak and trough serum levels of mnabs were estimated, to determine whether potency would be suf Measles Antibodies in Immunoglobulins JID 2006:194 (15 September) 781

2 ficient to maintain levels at or above those associated with protection. MATERIALS AND METHODS IGIV products. A total of 166 lots of 7 licensed IGIV products manufactured between 1998 and 2003 were randomly selected from lots submitted by manufacturers and released by the FDA. They were evaluated for mnabs, by the test described below. IGIV preparations were reconstituted, if lyophilized, according to the manufacturer s specification and were stored at either 20 C or 70 C, to ensure product integrity. Plaque-reduction neutralization test (PRNT). The PRNT was performed as described elsewhere and used low-passage Edmonston measles virus [9]. Immunoglobulin samples were diluted in PBS to a concentration of 1% IgG, except where noted, and were tested, in parallel, with the current US Reference IG (lot 176, manufactured in 1990 from Source Plasma) and the Second International Standard, 66/202 [10]. Geometric mean titers (GMTs) were calculated as log 10 -transformed reciprocal titers of mnabs and are reported as untransformed titers standardized against the Second International Standard. A PRNT titer of 1:8 is equivalent to 8 miu/ml. Preparation of purified IgG monomer, Fab, and F(ab ) 2. A single IGIV lot was used to prepare IgG monomer and fragments. Protein concentrations were determined on the basis of absorbance readings at 280 nm, with an absorption coefficient of 1.4; purity was monitored by size-exclusion high-performance liquid chromatography (SE-HPLC). Samples diluted to a 1% IgG solution in running buffer (0.02 mol/l Tris-chloride, 0.15 mol/l sodium chloride, 5 mmol/l EDTA, 0.02% sodium azide [ph 7.0]) were filtered through a 0.22-mm membrane. SE-HPLC was performed by use of a TSKGel-G3000SW column ( cm, 10-mm pore size; Tosoh Bioscience) preceded by a guard column ( cm); the flow rate was 0.3 ml/min. Multiple samples of IGIV were fractionated by SE-HPLC, and peak fractions of purified IgG monomer were pooled and were buffer-exchanged into PBS (ph 7.0) by use of a Centricon YM-50 concentrator (Millipore). Fab was prepared as follows: IGIV was diluted to a 5% IgG solution in 80 mmol/l sodium acetate buffer (ph 5.5) containing 50 mmol/l l-cysteine and 1 mmol/l EDTA and was incubated with 0.25 units papain-agarose beads (P-4406; Sigma Chemical)/mg IgG for 8 h at 37 C [11]. Digestion was terminated by use of a final concentration of 50 mmol/l iodoacetamide for 30 min at room temperature prior to filtration through a 0.45-mm Millipore filter and dialysis overnight (Slide- A-Lyzer; Pierce) into PBS (ph 7.0). The Fab fragment was further purified by protein G Sepharose mediated adsorption, according to the manufacturer s instructions (Amersham Pharmacia Biotech). F(ab ) 2 was prepared as follows: IGIV was diluted to a 5% IgG solution in 100 mmol/l sodium citrate buffer (ph 3.2) and was digested according to the method of Jones and Landon [12], by use of 10 units pepsin-agarose beads (P-0609; Sigma)/ mg IgG for 10 h at 37 C. After diafiltration in PBS (ph 7.0), residual Fc fragments were removed by adjustment of the dialyzed mixture to ph 4.0 prior to concentration by use of a Centricon YM-50 concentrator. The mixture was buffer-exchanged into 20 mmol/l piperazine, 150 mmol/l NaCl (ph 6.0) (buffer A) and then was passed through a Q-Sepharose Fast Flow column (Amersham Pharmacia) with a 2-mL bed volume. The F(ab ) 2 and Fab fragments recovered in the column flow-through and the first 2 3 washes with buffer A were concentrated to 1 ml for further purification. Residual intact IgG and Fc were removed by protein G Sepharose mediated adsorption. The peak fractions of F(ab ) 2 were collected via SE- HPLC fractionation, to remove any residual Fab, and then were pooled and buffer-exchanged into PBS (ph 7.0). Fractionation of IGIV into IgG subclasses. IgG subclasses were fractionated from 1 lot of an IGIV product manufactured from recovered plasma, as described by Scharf et al. [13]. Each fraction was diluted to a concentration of 0.1% IgG and was assayed by measles-virus specific PRNT. Specific activity was expressed as the quantity (in miu/mg) of measles antibody IgG that decreased the plaque count by 50%. Distribution of IgG subclasses, as determined by EIA. A total of 23 randomly selected IGIV lots, a subset of the 166 lots studied, were assayed according to the procedure described by Mikolajczyk et al. [14], to determine the percentage of each IgG subclass that was present. Calculated estimates of serum levels of mnab after administration of IGIV. The serum level of mnab after 1 dose of IGIV was estimated as follows: (1) the mean mnab potency was determined product by product; (2) peak titers were calculated according to the formula N/40, where N is the total dose (in miu) of IGIV administered to achieve the mean mnab potency divided by body mass (in kg); (3) the serum concentration at equilibrium between intravascular and extravascular compartments was estimated to be 40% of the peak titer, or [0.40 (N/40)] or (0.01N); and (4) the trough level was estimated to be one-half the level at equilibrium or (0.005N). Additional assumptions were (1) that the serum level of measles antibody, as determined by PRNT, prior to administration of IGIV was!4 miu/ml and (2) that the dose administered was 400 mg/kg body mass and that body mass was assumed to be 10 kg. The halflife of 1 dose of IGIV was estimated to be 22 days [15]. Statistical analysis. Statistical analysis was performed by use of unpaired Student s t test, and the results are expressed as the mean standard error of the mean (SEM). Results for which the P value was!.05 were considered to be significant. 782 JID 2006:194 (15 September) Audet et al.

3 Table 1. Summary of manufacturing procedures and lots tested, for 7 Immune Globulin Intravenous (Human) products. Lots tested, no. Year of manufacture Product Manufacturing procedures a Total A Cohn-Oncley (cold-ethanol) fractionation, PEG precipitation, ion-exchange chromatography, solvent/detergent treatment B Cohn-Oncley (cold-ethanol) fractionation, pasteurization C Cohn-Oncley (cold-ethanol) fractionation, ion-exchange chromatography, solvent/detergent treatment D Modified cold ethanol fractionation, PEG precipitation, ion-exchange chromatography, hydrolase incubation E Cohn-Oncley (cold-ethanol) fractionation, ion-exchange chromatography, solvent/detergent treatment F Cohn-Oncley (cold-ethanol) fractionation, solvent/detergent treatment, low-ph incubation G Modified cold ethanol fractionation, low-ph incubation with trace pepsin, with or without nanofiltration Total a Information is based on package inserts and includes virus-inactivation and -removal steps. PEG, polyethylene glycol.

4 RESULTS IGIV products may be manufactured from either recovered plasma or Source Plasma, and manufacture may include steps unique to specific products (table 1). It is generally understood that the demographic characteristics of the 2 donor populations are different. When the mnab GMTs for all IGIV lots tested were evaluated on the basis of year of manufacture, there was a trend toward decreasing anti-measles potency (1) in IGIV lots from recovered plasma that were manufactured between 2000 and 2002, compared with lots from recovered plasma that were manufactured in 1999, and (2) in IGIV lots from Source Plasma that were manufactured between 1999 and 2002, compared with lots from Source Plasma that were manufactured in 1998 (figure 1). The overall GMTs for lots manufactured from Source Plasma in 2002 and 2003 were significantly lower than the GMT for US Reference IG, which had an anti-measles potency of 2540 miu/ml at 1% IgG (or 41,910 miu/ml at 16.5% IgG) when assayed by PRNT. Interestingly, for 3 of the 6 years in which tests were conducted, the GMTs of IGIV lots derived from Source Plasma were significantly lower than those of IGIV lots derived from recovered plasma (figure 1). The mnab activities of IGIV lots were also evaluated product-by-product on the basis of year of manufacture and were compared with those of US Reference IG (figure 2). In this analysis, 6 of 7 IGIV products tested showed a decrease in antimeasles GMTs for lots manufactured in 2001 and 2002, compared with anti-measles GMTs for lots manufactured in Products C and G showed significant decreases in the antimeasles potency of IGIV lots manufactured in 2002, and product B showed a significant decrease in the anti-measles potency of IGIV lots manufactured in 2000, compared with the antimeasles potency of US Reference IG. In contrast, some products had significantly higher mnab potencies in IGIV lots manufactured in 1998 (products A and C) and in 1999 (products A and G). For all years of manufacture except 2001, product E had anti-measles GMTs that were significantly higher than that for US Reference IG. For no years of manufacture were the mnab GMTs for product F significantly different from that for US Reference IG. The most consistent finding was seen with product D, which had, in all 6 years considered, (1) anti-measles potency that was significantly lower than that of US Reference IG and (2) levels of mnab that were the lowest among the 7 IGIV products tested. We assessed whether the type of starting plasma, subclass distribution, and/or antibody fragmentation might alter the anti-measles potency of the final product. First, we examined whether the plasma used as the raw material for manufacture could affect levels of mnab, because Source Plasma is used in the manufacture of product D (table 2). We considered this possibility because the level of mnab in product E, which is derived from recovered plasma, was significantly higher than that in product C, which is derived from Source Plasma (GMT Figure 1. Potency of measles-virus neutralizing antibodies, in 7 Immune Globulin Intravenous (Human) (IGIV) products manufactured between 1998 and IGIV lots were diluted to a concentration of 1% IgG and then were tested in a measles-virus specific plaque-reduction neutralization test. Bars indicate geometric mean titers (GMTs) SEMs for IGIV products manufactured from Source Plasma (white bars) and for IGIV products manufactured from recovered plasma (shaded bars). * P!.05, compared with US Reference IG (US Ref IG); ** P!.05, for comparison of the 2 donor groups. 784 JID 2006:194 (15 September) Audet et al.

5 Figure 2. Potency of Immune Globulin Intravenous (Human) (IGIV) products, by year of manufacture, as determined by measles-virus specific plaquereduction neutralization test after dilution to a concentration of 1% IgG. Bars indicate yearly geometric mean titers (GMTs) SEMs for all products tested. * P!.05, for comparison with US Reference IG (US Ref IG). of product E and GMT of product C, miu/ml and miu/ml, respectively [ P!.05]), even though an identical procedure is used to manufacture both products. Overall, the levels of mnab in IGIV lots manufactured from recovered plasma (i.e., products E and G) were significantly higher than those of IGIV lots manufactured from Source Plasma (GMT SEM for 45 lots manufactured from recovered plasma and for 121 lots manufactured from Source Plasma, miu/ml and miu/ml, respectively [ P!.05]). Even when product D lots were excluded, the levels of mnab in lots manufactured from recovered plasma were still significantly higher than those in lots manufactured from Source Plasma (GMT SEM for 45 lots manufactured from recovered plasma and for 102 lots manufactured from Source Plasma, miu/ml and miu/ml, respectively [ P!.05]), suggesting that differences in product antimeasles potency may reflect the type of starting plasma used for manufacture. After IgG-subclass fractionation, the majority of mnab activity was found in the IgG1+IgG4 fraction (figure 3A); because IgG4 constituted 4% of total immunoglobulin, the majority of mnab activity in this fraction was most likely of type IgG1. The mnab activity of IgG3 was higher than that of IgG2, but both subclasses contributed only a small portion of the overall total mnab activity. The specific activities for mnabs for unfractionated IgG and of the IgG1+IgG4, IgG2, and IgG3 fractions were 239, 337, 39, and 68 miu/mg, respectively. Of the 7 IGIV products assayed, product D had the lowest level of IgG1 and was almost devoid of IgG3 (table 3); these low levels of both IgG1 and IgG3 could also help to explain the diminished mnab activity of this product. Similarly, the levels of IgG1 and IgG3 in product G (which also undergoes a protease treatment, albeit via a different method) were modestly below average (table 3), although its anti-measles potency was relatively high when compared with those of most other IGIV products (table 2). When the relative mnab activities of intact IgG monomer and of the F(ab ) 2 and Fab fragments were determined, on an equimolar basis, by PRNT, purified F(ab ) 2 exhibited 77% of the mnab activity seen in the IgG monomer, whereas the purified Fab exhibited only 20% of that same activity (figure 3B). The serum level of mnab theoretically achievable after 1 infusion of IGIV was also calculated. After a 400-mg/kg dose, all IGIV products can be expected to achieve serum levels 120 miu/ml (because mnab levels of 1:120 have been associated with protection in healthy vaccinees [16]) for 1 month after administration of IGIV (table 4). When each lot was considered individually, only 1 (product B) of 166 IGIV lots tested did not achieve, when given at a dose of 400 mg/kg, a serum level of mnab that was 120 miu/ml for the entire month after infusion, even though the trough titer estimated for this lot is 111 miu/ml (data not shown). On the basis of similar calculations, the theoretical minimum mnab potency, in IGIV at 1% IgG, that is needed to maintain serum levels at 120 miu/ ml for 1 month after a single 400-mg/kg dose is estimated to be 600 miu/ml. Measles Antibodies in Immunoglobulins JID 2006:194 (15 September) 785

6 Table 2. Summary of measles-virus neutralizing antibodies in 7 Immune Globulin Intravenous (Human) (IGIV) products manufactured between 1998 and Product Plasma type a GMT SEM b A Source ( n p 35) B Source ( n p 24) C Source ( n p 23) D Source ( n p 19) E Recovered c ( n p 22) F Source ( n p 20) G Recovered ( n p 23) US Reference IG d Source lower than those in products manufactured from recovered plasma. In addition, we believe that our findings reflect a specific change in anti-measles titers, because in-house assessment of several other antibody levels has not revealed declining trends during the same time period (data not shown). Intriguingly, there was an apparent increase, which we cannot fully explain, in the anti-measles potency of some IGIV products manufactured in The relative numbers of newly eligible young blood donors who have immunity due to natural infection could have increased because of measles outbreaks a Source Plasma is obtained from plasmapheresis donors; recovered plasma is from whole-blood collections. b Data in parentheses indicate total no. of lots tested. IGIV products and US Reference IG were diluted to a concentration of 1% IgG and were assayed by plaque-reduction neutralization test. GMT, geometric mean titer. c P!.05, for comparison of product E and product C, for comparison of products E+G and A+B+C+D+F, and for comparison when product D is excluded from the latter comparison. d 16.5% protein solution. The lot was tested in 37 assays. DISCUSSION Between 1995 and 2003 there was a steady increase in the proportion of recovered-plasma donors who had been born after 1956, with 49.9% of donors coming from this birth cohort in 1995 versus 57.7% in 2003 ([17] and E. Notari, personal communication). The percentage of Source Plasma donors born after 1956 was reported to be 58% in 1987 [18] and increased to 90% in 2003 (M. Gustafson, personal communication). As the proportion of donors with vaccine-induced immunity rises, one can expect a concomitant decline in mnabs in products derived from their plasma. An apparent decrease in mnab in IGIV lots manufactured between 1999 and 2002 is consistent with an increase in the proportion of recovered-plasma and Source Plasma donors born after 1956; the lower mnab potency of lots manufactured during this period may reflect an increase in donors who are more likely to have vaccine-induced measles immunity. Without additional information, we cannot prove that the decrease in anti-measles titers is the result of changes in donor demographic characteristics and/or measles epidemiology; nevertheless, we can speculate that the progressive shift in birth year will continue to be associated with an increase in the proportion of donors with vaccine-induced measles immunity and that this may result in a concomitant decrease in the anti-measles potency of products manufactured from these donors plasma. The lower anti-measles potencies seen in products derived from Source Plasma, compared with those in products derived from recovered plasma, also supports this hypothesis, because Source Plasma donors tend to be younger than recovered-plasma donors. This may explain why, during the same time period, the levels of anti-measles antibodies in products manufactured from Source Plasma are Figure 3. A, Potency of measles-virus neutralizing antibodies, by IgG subclasses. Bars indicate geometric mean titers (GMTs) SEMs for unfractionated IgG and isolated subclasses, after dilution to a concentration of 0.1% IgG. B, Neutralizing activity of fragments, tested at an initial concentration of 8.8 mmol/l. The apparent molecular weights used for calculation were 150,000, 100,000, and 45,000 for IgG monomer, F(ab ) 2, and Fab, respectively. Bars indicate the mean value of 2 assays. 786 JID 2006:194 (15 September) Audet et al.

7 Table 3. Distribution of IgG subclasses in 7 Immune Globulin Intravenous (Human) (IGIV) products and US Reference IG. Product a Distribution, mean SEM, % IgG1 IgG2 IgG3 IgG4 A( n p 3) B( n p 3) C( n p 3) D( n p 4) E( n p 4) F( n p 3) G( n p 3) US Reference IG a Data in parentheses are no. of IGIV lots assayed to determine distribution of IgG subclasses, as described in Materials and Methods. that occurred in the United States in the 1980s and thereby could have influenced anti-measles potency in lots manufactured in 2003 [19, 20]. Likewise, certain manufacturing procedures that include protease treatments and other downstream procedures could differentially affect specific IgG subclasses and thus potentially alter the anti-measles potency of the final product. Scharf et al. [13] showed that the dominant neutralizing IgG subclass for HIV-1 in an experimentally produced HIV-specific immune globulin was IgG3, a subclass known to be extremely sensitive to protease treatment. Although previous work had shown that measles-virus binding antibodies are primarily IgG1, the subclass profile of the functional (i.e., neutralizing) mnab response in humans had been unknown [21, 22]. The present report is the first to document the measles-virus neutralizing activity of each IgG subclass, and it demonstrates that IgG1 has the highest specific measles-virus neutralizing activity, with only a minor additional contribution from IgG3. It appears that the loss of both IgG1 and IgG3 in 1 of the IGIV products that we tested may be responsible for the latter s relatively low measles-virus neutralizing activity. Endogenous plasma proteases may copurify with IgG during manufacture and thus result in IgG fragmentation and diminished measles-virus neutralizing activity in the final container. The results of the present study indicate that measles neutralization requires bivalent antibodies, such as intact IgG and F(ab ) 2, whereas the monovalent Fab was observed to be less active on a molar basis; however, fragmentation is usually not Table 4. Estimated serum levels of measles-virus neutralizing antibodies (mnabs) after administration of Immune Globulin Intravenous (Human) (IGIV) manufactured between 1998 and Product Mean mnabs, miu/ml a Total mnabs per dose, miu b Serum levels of mnabs, miu/ml c Administration (day 0) Equilibrium (days 4 5) Trough (days 26 28) d A ,123, B , C ,030, D , E ,436, F , G ,054, US Reference IG , e 105 e Minimum potency f , a Assayed at 1% IgG concentration; for details, see table 2. b Administered dose was 400 mg/kg except in the case of US Reference IG, for which it was 0.5 ml/kg; estimated weight of each pediatric recipient was 10 kg. c Based on the formula described in Materials and Methods. d Half-life of IgG was assumed to be 22 days. e Average values, which can vary slightly depending on the rate and pattern of absorption. f Theoretical minimum is the minimum potency needed for an IGIV product to achieve a serum level of 120 miu/ml, for 1 month after a single 400-mg/kg dose of IGIV. Measles Antibodies in Immunoglobulins JID 2006:194 (15 September) 787

8 a problem, because endogenous proteases are inactivated by low-ph treatment or by freeze-drying or are removed by chromatography methods used to purify some immunoglobulin products. Although only IG is approved for measles prophylaxis, IGIV is expected to prevent a variety of bacterial and viral infections in patients with PIDD. All IGIV lots evaluated in the present study met the lot-release specifications set by the FDA. Although mnab titers 1:120 have been associated with protection against severe measles disease in healthy vaccinated persons [16], the precise serum level of measles antibody necessary to prevent infection in immunodeficient measles-susceptible individuals is not known. IG was originally licensed for measles prophylaxis before the isolation of measles virus, and protective doses were determined clinically on the basis of prevention of the disease after IG treatment during measles outbreaks [23, 24]. The absence of fulminant measles infection in individuals with PIDD who are treated with either IG or IGIV provides additional indirect evidence that these products have measles antibody sufficient to provide protection; however, studies to document the pharmacokinetics of measles antibodies after injection or infusion in this population have not been conducted. More recently, it has been suggested that individuals who receive immunoglobulins containing 140 IU of measles antibodies in the total administered dose will be protected from measles disease [25]; unfortunately, that study did not report the measles immune status before treatment and did not measure either peak or trough levels of measles antibody after infusion. Likewise, in a patient cohort undergoing bone-marrow stem-cell transplantation, measles-antibody titers were measured and were found to be 900 miu/ml after each of 2 infusions of IGIV and were maintained at or above this level for 21 days after the last infusion [26]; however, these patients had preexisting measles immunity, and the reported mnab levels after immunoglobulin infusion may not reflect the levels achievable in measles-naive individuals with PIDD. A single dose of IG with the minimum anti-measles potency (i.e., 0.6 times that of the US Reference IG, lot 176) would contain 25,000 miu/ml of measles antibody and, at the maximum dose of 15 ml, would provide 380,000 miu of measles antibody, or a minimum of 5000 miu/kg for a 70-kg adult. In contrast, IGIV having the minimum anti-measles potency would, when given at a dose of 400 mg/kg, provide that 70-kg individual with 4,270,000 miu of measles antibody, or 61,000 miu/kg. The minimum quantity of measles antibody necessary for measles prophylaxis in immune-deficient individuals is not known; therefore, the clinical benefits of one product over the other cannot be assessed. It is obvious, however, that, in adults, the per-dose measles antibody provided by IGIV products could possibly be greater than that provided by a corresponding maximum dose of IG, if both products are at or near the current minimum antimeasles potency. Our calculations predict that all IGIV products (except for 1 product B lot) would provide measles antibody sufficient to maintain serum levels of 120 miu/ml between monthly infusions. Also, calculations using a half-life of 22 days for IgG may over- or underestimate the potential longevity of measles antibody in some patients; this variation must be considered in any estimation of the protection offered against measles infection. The level of mnabs in plasma pools will likely diminish as the number of vaccinated donors increases and as circulation of wild-type measles virus is curtailed by herd immunity. We believe that a scientifically sound approach for addressing the use of immunoglobulin therapy for measles prophylaxis in the vaccine era would be to determine both (1) the minimum titer associated with protection against measles infection in immune-deficient patients and (2) the dose of IGIV necessary to achieve this titer. This approach should be discussed in the broader context of measles epidemiology, with data provided from pharmacokinetic studies of immune-deficient patients that measure how actual (rather than theoretical) trough levels of measles antibodies relate to the amount infused. Acknowledgments We thank Ed Notari (American Red Cross) and Mary Gustafson (Plasma Protein Therapeutics Association), for providing donor demographic data; Dr. Timothy Forsey (National Institute for Biological Standards and Control, United Kingdom), for providing the Second International Standard (66/202); and Drs. John S. Finlayson and Joe Kutza, for their critical review of the manuscript. References 1. Immune Deficiency Foundation. Primary immune deficiency diseases in America: the first national survey of patients and specialists. Available at: Accessed 9 August Knezevic-Maramica I, Kruskall MS. Intravenous immune globulins: an update for clinicians. Transfusion 2003; 43: Office of Genomics and Disease Prevention, Centers for Disease Control and Prevention. Applying genetics and public health strategies to primary immunodeficiency diseases. Available at: genomics/info/conference/pi/pimeeting2.htm. Accessed 9 August Siegel J. 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