Murine B Cells Regulate Serum IgE Levels in a CD23-Dependent Manner
The manifestations of allergic disorders are closely tied to the biologic effects of IgE activation with Ag. In immediate hypersensitivity reactions, IgE effector function requires prior binding to innate immune cells, primarily mast cells and basophils, with the blood acting as a reservoir for unbound IgE. As the severity of allergic disease is proportional to the size of this unbound IgE pool, we hypothesized that cellular mechanisms exist to limit the size and/or enhance the clearance of free IgE molecules. We examined this in mice by engineering a reporter IgE molecule that allowed us to track the fate of IgE molecules in vivo. The absence of FcεRI-expressing cells did not affect serum IgE levels, but B cells regulated serum IgE by controlling the size of the free IgE pool. B cells captured IgE by direct binding to the low-affinity IgE receptor, CD23. These data indicate a mechanism regulating serum IgE and additionally clarify the role of CD23 in this process.
The Journal of Immunology
Murine B Cells Regulate Serum IgE Levels in a
Laurence E. Cheng,* Zhi-En Wang,
and Richard M. Locksley
The manifestations of allergic disorders are closely tied to the biologic effects of IgE activation with Ag. In immediate hypersen-
sitivity reactions, IgE effector function requires prior binding to innate immune cells, primarily mast cells and basophils, with the
blood acting as a reservoir for unbound IgE. As the severity of allergic disease is proportional to the size of this unbound IgE pool, we
hypothesized that cellular mechanisms exist to limit the size and/or enhance the clearance of free IgE molecules. We examined this
in mice by engineering a reporter IgE molecule that allowed us to track the fate of IgE molecules in vivo. The absence of Fc«RI-
expressing cells did not affect serum IgE levels, but B cells regulated serum IgE by controlling the size of the free IgE pool. B cells
captured IgE by direct binding to the low-afﬁnity IgE receptor, CD23. These data indicate a mechanism regulating serum IgE and
additionally clarify the role of CD23 in this process. The Journal of Immunology, 2010, 185: 5040–5047.
he clinical manifestations of allergic disease result from
the effects of a complex network of cells and effector mole-
cules (1). Among these, IgE plays a central role in medi-
ating allergic disease. Acute allergic reactions, such as anaphylaxis,
are primarily the result of IgE-mediated processes, although IgG
can mediate similar effects (2–4). IgE also contributes to chronic
allergic diseases, including asthma and atopic dermatitis. In these
diseases, the serum levels of total or speciﬁc IgE correlate closely
with disease severity (5, 6). Therefore, the regulation of IgE pro-
duction and/or clearance is central to allergic disorders.
Among the soluble Ig isotypes, IgE exhibits several unique
properties. First, the most prominent effects of IgE effector func-
tion, as seen in type I hypersensitivity reactions, require prior
binding to the FcεRI. FcεRI is a heterotetrameric complex ex-
pressed primarily on the surface of mast cells and basophils. The
complex is composed of an a-chain (which binds IgE), a b-chain
(which ampliﬁes signaling), and a dimer of g-chains (which is the
primary signaling component) (7). Loading of IgE onto mast cells
and basophils enables these cells to recognize and respond to
speciﬁc Ags with immediate release of preformed molecules,
synthesis of lipid mediators, and cytokine production (8–10). In
humans, APCs, including Langerhans cells, express a trimeric
form of the FcεRI (ag
), which play a role in Ag processing (11).
cells are found throughout the body, mast cells
are the most abundant, with a cellular distribution weighted to-
ward peripheral tissues, including the skin as well as the gastro-
intestinal and respiratory tracts (12).
A low-afﬁnity IgE receptor (CD23 or FcεRII) is also present and
exists as membrane-bound trimers as well as soluble monomers
and oligomers (13, 14). In mice, CD23 is expressed primarily on
B cells, although follicular dendritic cell expression has also been
described (15, 16). Human CD23 has a broader expression pattern
with the existence of two CD23 isoforms (17). Membrane-bound
CD23 has been shown to both enhance and diminish IgE pro-
duction by B cells, to facilit ate the clearance of IgE immune
complexes, and to mediate transport of IgE across the gut epithe-
lium (18, 19). The soluble form has an order of magnitude lower
afﬁnity for IgE than does the membrane-bound form and may
indirectly enhance IgE production (9). CD23 also has a broader
ligand-binding proﬁle than does FcεRI, which may account for
the disparate effects of CD23 on IgE production (20, 21).
The amount of free IgE available for binding to receptors is also
unusual. Free IgE is found primarily in the blood and is the scarcest
serum Ig. In normal children and adults, IgE is found at 100- to
10,000-fold lower levels than IgG (8, 9). This paucity of IgE
reﬂects the rapid clearance of free IgE from the blood. Whereas
murine IgA, IgM, and IgG have serum half-lives on the order of
a day to weeks, the half-life of free IgE is 5–12 h (22, 23). The
half-life of human IgE in normal individuals is longer than mice
(∼2 d), but it is still the shortest among soluble Igs (24).
The rapid clearance of unbound IgE from the serum is in part
related to the catabolism of serum IgE (24, 25). However, other
mechanisms may also exist. In severely atopic individuals, IgE
levels increase up to 1000-fold, and the rate of serum IgE clear-
ance in these individuals is inversely related to this rise in serum
IgE (24, 25). This contrasts with IgM, IgG, and IgA, for which the
clearance rate increases proportionally with increases in serum Ig
levels. These data suggest that a saturable mechanism may exist to
regulate serum IgE levels and/or alter IgE clearance. Given the
presence of at least two IgE receptors and the relative abundance
of IgE-binding cells, we hypothesized that a cellular mechanism
may mediate IgE clearance. Our data indicate that CD23 expres-
sion on B cells, rather than FcεRI
mast cells or basophils, reg-
ulates free serum IgE levels. These data reveal a cellular mech-
anism for monomeric Ab clearance and suggest an additional
function for circulating B cells.
*Department of Pediatrics,
Department of Medicine,
Department of Microbiology
and Immunology, and
Howard Hughes Medical Institute, University of California,
San Francisco, San Francisco, CA 94143
Received for publication June 10, 2010. Accepted for publication August 29, 2010.
This work was supported in part by the Howard Hughes Medical Institute, National
Institutes of Health Grants AI026918 and AI078869 (to R.M.L.), a T32 National
Institute of Child Health and Human Development Institutional Training Grant
(HD044331), an American Academy of Allergy, Asthma and Immunology/Glaxo
SmithKline Career Development Award, and the A.P. Giannini Medical Research
Foundation (to L.E.C.).
Address correspondence and reprint requests to Dr. Richard M. Locksley, University
of California, San Francisco, 513 Parnassus Avenue, Box 0795, San Francisco, CA
94143. E-mail address: email@example.com
The online version of this article contains supplemental mate rial.
Abbreviations used in this paper: HA, hemagglutinin; HA-Fcε, HA-tagged Fcε; MFI,
mean ﬂuorescence intensity; rPCA, reverse passive cutaneous anaphylaxis; TNP,
Copyright Ó 2010 by The American Association of Immunologists, Inc. 0022-1767/10/$16.00
Materials and Methods
BALB/c, C57BL/6, and C57BL/6 FcεRI
mice were from The Jackson
Laboratory (Bar Harbor, ME) (26). mMT mice were from The Jackson
Laboratory and were backcrossed to BALB/c mice for 10 generations (27).
mice have been described (28, 29). Mast cell-
mice on a C57BL/6 background were provided by
G. Caughey (University of California, San Francisco) (30). 4get BALB/c
and 4get Rag2
mice have been described (27). Mice were housed in
speciﬁc pathogen-free facilities. Experimental mice were 8–12 wk old.
Animal use was governed by and in accordance with approved protocols
overseen by the Laboratory Animal Resource Center and Institutional
Animal Care and Use Committee at the University of California, San
Abs and ﬂow cytometry
We used the following Abs and clones: anti–c-kit (2B8; BioLegend, San
Diego, CA), anti-CD49b (DX5; eBioscience, San Diego, CA), anti-mouse
IgE (23G3; SouthernBiotech, Birmingham, AL), B220 (RA3-6B2; BD
Biosciences, San Jose, CA), and CD19 (1D3; BD Biosciences). Hemag-
glutinin (HA) tags were detected with a mAb conjugate (GG8-1F3.3.1;
Miltenyi Biotec, Auburn, CA). We used an LSRII ﬂow cytometer (BD
Biosciences) for cell acquisition and analysis. For intracellular HA staining,
ﬁxed splenocytes were treated for 5 min with 0.5 U/ml Liberase CI (Roche,
Indianapolis, IN) at 37˚C in an orbital shaker. The reaction was immediately
quenched with PBS containing 2% FCS. Cells were then permeabilized in
PBS supplemented with 2% FCS and 0.5% saponin. We performed post-
acquisition analysis using FlowJo software (Tree Star, Ashland, OR).
Reverse passive cutaneous anaphylaxis
We performed local hypersensitivity challenges following passive sensi-
tization similar to published protocols (31). In brief, we sensitized mice
with varying amounts of anti-trinitrophenol (TNP) IgE Ab (C38-2) of the
allotype (BD Biosciences). Two days later, we loaded the mice with
200 ml of 1% Evans blue dye in PBS and challenged the mice with a 1-mg
intradermal ear injection of TNP
-OVA (Biosearch Technologies, Novato,
CA) in PBS. One hour after challenge, we isolated the ears, weighed the
tissue, and placed the tissue in formamide overnight at 55˚C. We then
ﬁltered the supernatant and measured the absorbance at 620 nm. The ab-
sorbance was corrected for background values and then divided by the
weight of the tissue.
Construction of HA-tagged Fc«
We isolated IgE cDNA from an IgE-secreting hybridoma, IGEL4.1a (provided
by M. Wabl, University of California, San Francisco), which encodes the a
allotype of the IgE H chain sequence, Cε1–Cε4, using RT-PCR. This fragment
was subcloned using Zero Blunt TOPO (Invitrogen, Carlsbad, CA) and
the sequence was veriﬁed. The Cε2–Cε4 cDNA, which includes two disulﬁde
bonds required for homodimerization, was ampliﬁed using the following
primer pair: 59-AGATCTGTTCGACCTGTCAACATCAC-39 and 59-GTTC-
GTCGACGGGCCCG-39. We cloned this amplicon in-frame using a BglII/
SalI digestion into an expression vector derived from pcDNA-3, resulting in
the addition of a V
signal sequence and HA epitope tag on the N terminus
of the protein. To enhance expression of the HA-tagged Fcε (HA-Fcε), we
subcloned the cDNA to an expression vector containing the elongation factor-
1a promoter, pShooter (Invitrogen). We then transfected the hybridoma fusion
partner SpAg14 by electroporation with 10 mg of ScaI-linearized plasmid
DNA. We selected stable clones with 2 mg/ml G418 and veriﬁed protein
production by ELISA.
For native IgE and HA-Fcε capture, we used anti-IgE clone RME-1
(eBioscience). IgE detection was performed using biotinylated anti-IgE
(EM95), followed by streptavidin-conjugated alkaline phosphatase (BD
Biosciences). HA detection was performed using an anti-HA HRP con-
jugate (Miltenyi Biotec).
Similar to published protocols (32, 33), we infused 100 mg of B3B4
(BioLegend) or isotype control rat IgG2a Ab into the tail vein, followed by
the indicated treatments.
IgE levels correlate with severity of the allergic response
Clinical data suggest a direct correlation between the amount of
Ag-speciﬁc serum IgE and the degree of local hypersensitivity.
Therefore, we used a reverse passive cutaneous anaphylaxis (rPCA)
assay to determine the extent of IgE loading on mast cells in
peripheral tissues following i.v. infusion with increasing amounts
of Ag-speciﬁc IgE. We infused BALB/c mice with 0.25–5 mgof
TNP-speciﬁc IgE, and 2 d later we administered Evans blue dye
by i.v. injection, followed by a challenge with 1 mg of TNP-OVA
into the ear by intradermal injection. One hour after challenge, we
harvested ear tissue and extracted Evans blue dye. The rPCA
procedure caused some dye to extravasate even in the absence of
IgE. However, the degree of dye extravasation was directly pro-
portional to the amount of IgE, with a maximal response at the
highest dose of IgE administered (Supplemental Fig. 1). Thus,
these data indicate that the amount of serum IgE is proportional to
the subsequent local hypersensitivity response.
Construction and characterization of a surrogate IgE molecule
We next sought to develop a system to track the disposition of
serum IgE in IgE-sufﬁcient animals. To accomplish this, we
engineered HA-Fcε by fusing an HA tag N-terminal to the Cε2–
Cε4 domains of the IgE H chain (Fig. 1A). The crystal structure of
the Fcε was based on a similar molecule, and prior chimeric
molecules have been similarly engineered (34, 35). Although HA-
Fcε would be predicted to have a similar structure to native IgE
molecules, we wanted to ensure that HA-Fcε displayed similar
clearance kinetics as native IgE. Thus, we infused IgE-deﬁcient
mice (which lack detectable serum IgE and lack cell
surface IgE on basophils or mast cells) with 1 mg of IgE or HA-
Fcε. We serially bled the mice and assayed for serum IgE levels
at 0, 1, 7.5, and 24 h (Fig. 1B). Just prior to the infusion, we found
no detectable IgE in these mice. IgE levels were highest at 1 h
postinfusion and rapidly declined by 7.5 h. By 24 h, no serum IgE
was detectable in either group. The levels of serum IgE at each
time point were nearly identical between the two groups.
We next compared the capacity of HA-Fcε and native IgE to
bind to FcεRI
basophils. To facilitate the detection of FcεRI
splenic basophils, we used Rag2
mice on a 4get background
(27). These mice (4getRag2
) contain the coding sequence of
the enhanced GFP under the translational control of an internal
ribosomal entry site at the 39 end of the last exon of the IL-4 gene.
Basophils, mast cells, and eosinophils from such mice constitu-
tively express GFP (36). To examine binding of HA-Fcε to
basophils, IgE-deﬁcient 4getRag2
mice received 1 mg of HA-
Fcε or control IgE by tail vein injection. Twenty-four hours later,
we analyzed splenic basophils for surface IgE levels and the
presence of HA-Fcε. As expected in the absence of serum IgE,
splenic basophils from 4getRag2
mice showed no detectable
IgE staining at baseline. After infusion, IgE was readily detected
on splenic basophils (Fig. 1C), with similar mean ﬂuorescence
intensity (MFI) of IgE staining in IgE- and HA-Fcε–infused ani-
mals. In contrast, only splenic basophils from HA-Fcε–infused
animals had detectable HA staining (Fig. 1C,1D).
We next addressed whether HA-Fcε was capable of binding to
basophils and mast cells in IgE-sufﬁcient hosts. Additionally, we
examined the amount of HA-Fcε sufﬁcient to uniformly load both
basophils and peripheral mast cells with IgE. Therefore, we ad-
ministered increasing doses of HA-Fcε from 0.1 to 10 mg to IgE-
sufﬁcient 4get mice on a BALB/c background. Twenty-four hours
after infusion, both basophils and mast cells readily accumulated
cell surface HA-Fcε, and basophils showed detectable staining
The Journal of Immunology 5041
even at the lowest infusion dose (Fig. 2). Presumably, peritoneal
mast cells required higher infusion doses owing to differential
delivery of HA-Fcε in blood as compared with the peritoneum.
At the 2.5-mg dose, both peritoneal mast cells and basophils
demonstrated uniform acquisition of HA-Fcε. Taken together,
HA-Fcε demonstrates similar kinetic and biochemical properties
to native IgE.
B cells control the set point for serum IgE levels
Given the relative abundance of IgE receptor-bearing cells, we
hypothesized a cellular control mechanism for serum IgE levels.
cells have previously been shown to have little
bearing on the regulation of serum IgE (26), we wanted to verify
these ﬁndings using our reporter IgE molecule. Therefore, we
administered 2.5 mg of HA-Fcε to FcerI
mice or wild-type
C57BL/6 controls and performed serial bleeds at 1, 3, 7.5, and
∼24 h postinfusion (Fig. 3A). The peak HA-Fcε level in the blood
at 1 h was nearly identical between the two groups of mice, and
there were no signiﬁcant differences between the two groups at the
remaining time points. Experiments in mast cell-deﬁcient Sash
) yielded similar results (data not shown).
We next examined whether B cells, which, in mice, are the
primary CD23-expressing cells, contribute to regulation of serum
IgE (8, 13). B cell-deﬁcient mMT
mice received 2.5 mg of HA-
Fcε by tail vein injection. Serial assessment of HA-Fcε levels
revealed a 2-fold increase in the peak IgE level at 1 h compared
with wild-type controls (Fig. 3B). Whereas B cells affected the
serum IgE level, the absence of B cells had no impact on the
overall clearance kinetics of HA-Fcε, as B cell-deﬁcient and
-sufﬁcient mice had similar half-lives of 5.9 and 5.2 h, re-
spectively. We also observed similar results in Rag2
not shown). To ensure that the differences were solely due to the
FIGURE 1. Construction and characterization of HA-Fcε. A, Schematic of the HA-Fcε construct. The Cε2–Cε4 domains of the IgE H chain are po-
sitioned C-terminal of a V
signal sequence and the HA epitope tag. B, Groups of two to three IgE-deﬁcient 4getRag2
mice received 1 mg of native IgE
(n) or HA-Fcε (:) at time 0. We then serially bled the mice to follow the fate of infused IgE. Error bars represent the SEM. C and D, 4getRag2
received 1 mg of native IgE (C) or HA-Fcε (D) by tail vein injection. Twenty-four hours later, we analyzed splenic basophils (GFP
) for the
presence of both total IgE and HA (solid black lines). The gray histograms indicate the staining of negative control IgE-deﬁcient mice. Data are repre-
sentative of at least three independent experiments.
FIGURE 2. Uptake of HA-Fcε by splenic basophils and peritoneal mast cells. We infused 4get BALB/c mice with increasing amounts of HA-Fcε as
listed at the top of the ﬁgure. Twenty-four hours later, we analyzed GFP
splenic basophils (A) or GFP
peritoneal mast cells (B) for
HA-Fcε using an anti-HA Ab. The percentage of HA-Fcε
cells is depicted with each histogram. Data are representative of two independent experiments.
5042 B CELL REGULATION OF SERUM IgE
absence of B cells (rather than IgE), we performed similar
experiments in Stat6- and IL4/13-deﬁcient animals. Both of these
animals have B cells but no serum IgE or detectable surface IgE
cells. Despite the absence of IgE, wild-type, Stat6
mice had overlapping peak serum IgE levels and
clearance kinetics (Fig. 3C). Taken together, these data suggested
a role for B cells in controlling the serum IgE level but not the rate
of IgE clearance.
CD23 blockade is sufﬁcient to inhibit B cell regulation of IgE
We next examined whether CD23 expression on B cells was re-
sponsible for the B cell-dependent regulation of serum IgE levels.
CD23 is thought to have multiple functions on B cells, and the anti-
CD23 mAb B3B4 antagonizes IgE binding to CD23 (32). We
therefore administered 100 mg of B3B4 or isotype control Ab to
groups of mice and, 1 d later, examined clearance of a 2.5-mg HA-
Fcε challenge (Fig. 4A). At 1 h postinfusion, B3B4-treated mice
exhibited a 1.8-fold increase in serum IgE as compared with
isotype controls. This increase was similar to the differences seen
in B cell-deﬁcient animals as compared with controls. CD23
blockade also did not affect the overall clearance of HA-Fcε,as
the curves showed parallel declines in IgE with nearly complete
absence of HA-Fcε by 24 h in both groups of animals. Binding of
HA-Fcε to peritoneal mast cells similarly reﬂected the increased
serum IgE levels, as B3B4-treated mice demonstrated a 7-fold
increase in MFI on mast cells as compared with isotype controls
(Fig. 4B). The effect on splenic basophils was similar but more
modest (Fig. 4C).
Taken together, our data indicate that CD23 expression by B cells
regulates serum IgE levels. CD23 can, however, exist in both soluble
and membrane-bound forms, and our data do not preclude a role
for soluble CD23 in IgE clearance. To investigate whether B cells
directly bind IgE, we ﬁrst veriﬁed CD23 expression on splenic
B cells (Fig. 5A). Most B cells were CD23
, similar to published
data (37). We then examined IgE binding to B cells by infusing
BALB/c mice with 2.5 mg of HA-Fcε. Two or 24 h after infusion,
we generated single-cell suspensions and immediately ﬁxed the
cells with 4% paraformaldehyde, which enhanced detection of cell
surface HA-Fcε on B cells. We then analyzed CD19
for HA-Fcε capture by ﬂow cytometry. B cells from mice that had
received PBS demonstrated no HA staining 2 h after infusion. In
contrast, B cells from mice, which had received HA-Fcε 2 h before,
demonstrated uptake of HA-Fcε (Fig. 5B). This binding of HA-Fcε
was short-lived and directly proportional to the serum HA-Fcε
level, as no HA-Fcε was detectable on B cells 24 h after infusion.
To ensure that the ﬁxation did not result in nonspeciﬁc binding
to cells, we veriﬁed that neither CD4
T cells showed
HA-Fcε binding after ﬁxation (data not shown).
We next examined whether HA-Fcε binding to B cells was CD23
dependent. Groups of mice received pretreatment with B3B4 (or
isotype cont rol) foll owed by 2.5 mgofHA-Fcε the next day.
Administration of B3B4 completely blocked HA-Fcε binding t o
B cells 2 h after treatment wi th HA-Fcε (Fig. 5C).
Following binding to CD23, IgE molecules can be internalized
and degraded (38). Thus, we examined whether splenic B cells
internalized HA-Fcε after binding. BALB/c mice received either
PBS or HA-Fcε, and 2 h later, we harvested spleens and ﬁxed
single-cell suspensions as above. To differentiate between extra-
and intracellular HA-Fcε, the samples were split into two groups.
One group was stained for CD19, B220, and HA. A second group
was treated brieﬂy with Liberase CI to remove extracellular HA-
Fcε. As before, B cells had cell surface HA-Fcε 2 h after infusion
(Fig. 5D), and these molecules were almost completely stripped
FIGURE 3. B cells regulate serum IgE levels. A, Wild-type (n)orFcεRI
(:) mice received 2.5 mg of HA-Fcε by tail vein injection. We then analyzed
HA-Fcε levels at the indicated time points. B, We analyzed wild-type (n)ormMT
(:) animals for HA-Fcε clearance at the indicated time points.
C, Wild-type (n), Stat6
(:), or IL-4/13
(▼) mice received 2.5 mg of HA-Fcε and were analyzed at the indicated time points. Data are representative
of at least two independent experiments each. Wild-type and FcεRI
groups had three mice each. mMT and Stat6
groups had four mice each. The
group had two animals. Error bars represent the SEM. pp , 0.001; ppp , 0.007.
FIGURE 4. Anti-CD23 blockade increases peak serum IgE levels and enhances IgE loading. A, BALB/c mice received either 100 mg of B3B4 (:)or
isotype (n) control Ab by tail vein injection. One day later, the mice received 2.5 mg of HA-Fcε and were serially bled to examine HA-Fcε serum levels.
There were ﬁve mice in each group, with error bars representing the SEM. pp , 0.015; ppp , 0.005. B and C, The mice received B3B4 (solid black line) or
isotype (dashed line) as above and were given 0.5 mg of HA-Fcε the next day. Surface HA-Fcε capture was assessed on (B) peritoneal mast cells and (C)
splenic basophils 24 h later. The MFI is noted above the histograms. The shaded histograms represent the negative control. The data are representativeof
two independent experiments.
The Journal of Immunology 5043
off after brief exposure to Liberase CI. Subsequent permeabil-
ization of the cells led to an increase in HA staining. Taken to-
gether, these data demonstrate that IgE binds directly to B cells in
a CD23-dependent manner and that bound IgE is subsequently
CD23 blockade enhances IgE-mediated hypersensitivity
Because CD23 blockade increases the peak serum IgE level, we
next examined whether this increase leads to greater delivery of
IgE to tissue mast cells and enhanced local hypersensitivity re-
actions after Ag challenge. To address this, we blocked CD23 in
BALB/c mice with 100 mg of B3B4 on day 0. On day 1, the mice
received 0.5 mg of anti-TNP IgE by tail vein injection. We chose
this dose as it is on the linear portion of the dose–response curve
(Supplemental Fig. 1). On day 3, we administered Evans blue and
challenged the mice with 1 mg of TNP-OVA in the rPCA assay.
As compared with mice that lacked speciﬁc IgE, isotype control-
treated mice had a signiﬁcant increase in Evans blue dye extra-
vasation 1 h after challenge (Fig. 6). In the presence of B3B4, this
effect was further enhanced with a 40% increase in Evans blue
after challenge. These data indicate that blockade of CD23-
dependent IgE binding signiﬁcantly increases the severity of IgE-
CD23 blockade leads to a rapid increase in total serum IgE
Our data suggest that cell surface-bound IgE has a rapid turnover
(Fig. 5B), and we therefore expected blockade of CD23 to lead to
an increase in total serum IgE soon after administration. To ex-
amine this, we administered B3B4 (or isotype control) to groups
of BALB/c mice and performed serial bleeds 1, 4, and 7 d after
infusion to monitor total serum IgE levels. Consistent with a role
for CD23 in regulating serum IgE levels and rapid turnover of IgE
bound to B cells, the serum IgE level nearly doubled within 24 h
of CD23 blockade (Fig. 7). This effect began to wane by 4 d and
was nearly absent by 7 d postinfusion. These data indicate that
CD23 blockade not only has effects on passively infused IgE but
also the native IgE pool.
Our data reveal an unproven in vivo role for murine B cells in the
regulation of serum IgE levels. B cells act as a sink to eliminate
excess serum IgE by binding to IgE in a CD23-dependent manner.
FIGURE 5. B cells bind HA-Fcε in a CD23-
dependent manner. A, CD19
BALB/c mice were stained with B3B4 (solid
line) or isotype control (shaded histogram). B,
BALB/c mice received either PBS or 2.5 mgof
HA-Fcε. We harvested splenocytes 2 or 24 h
after HA-Fcε infusion and ﬁxed the cells in 4%
paraformaldehyde immediately. The time point
at harvest is indicated in each of the histograms.
We stained single-cell suspensions with CD19,
B220, and anti-HA Abs. The solid lines in the
each of the panels represent cell surface HA-Fcε
cells. The shaded histograms
represent staining on B cells from mice that re-
ceived PBS. C, Two hours after receiving 2.5 mg
of HA-Fcε, ﬁxed B cells from mice that had
received 100 mg of isotype control (left panel)or
B3B4 (right panel) 1 d prior were examined as
above. D, Two hours after receiving 2.5 mgof
HA-Fcε, ﬁxed B cells from mice were stained for
extracellular HA-Fcε (left panel) or intracellular
HA-Fcε (right panel). The middle panel depicts
extracellular HA-Fcε staining after collagenase
digestion. Shaded histograms represent B cells
from PBS-infused animals. Data are representa-
tive of at least two independent experiments.
FIGURE 6. CD23 blockade enhances local hypersensitivity reactions.
Mice received either 100 mg of isotype control or B3B4 Ab on day 0. On
day 1, the mice were loaded with 0.5 mg of anti-TNP IgE. On day 3, the
mice received 200 ml of 1% Evans blue dye by tail vein injection and then
a1-mg intradermal injection of TNP
-OVA into the ear. A non-IgE–loaded
group of mice served as a background control. Average values were de-
termined from the ears from two mice in the nonloaded group, three mice in
the isotype-treated group, and four mice from the B3B4-treated group. The
data are representative of two independent experiments. pp , 0.01.
5044 B CELL REGULATION OF SERUM IgE
Although B cells eliminated excess serum IgE, the overall rate of
serum IgE catabolism remained largely unchanged.
The lack of contribution by FcεRI
cells to the clearance of IgE
has previously been reported (26). These observations are partic-
ularly striking given the afﬁnity of the FcεRI for IgE (subnano-
molar range) (9). One possible explanation is that only a limited
number of unoccupied receptors exist on the surface of basophils
and mast cells. Although quantifying the numbers of unoccupied
receptors is difﬁcult (39), it was clear that both basophils and
mast cells had sufﬁcient unoccupied receptors to bind exogenous
IgE (Fig. 2). Indeed, the regulation of surface FcεRI is thought to
occur mainly through stabilization of unoccupied receptors after
IgE binding, with a dynamic balance between de novo FcεRI pro-
duction and endocytosis of unoccupied receptors (40). The number
of target cells available may also have been a factor. Basophils are
the least represented hematopoietic cell in the peripheral blood,
found at levels ∼10% of the numbers of B cells in normal mice and
humans. Furthermore, B cells are heavily represented in secondary
lymphoid organs such as the spleen and lymph nodes, while baso-
phils are more rarely observed in these tissues (41, 42). Basophils
undergo rapid turnover and could thereby regulate serum IgE lev-
els (43); however, these effects would need to be in an FcεRI-
independent manner. In contrast, mast cells are widely distributed
in peripheral tissues, including the skin, intestines, and respiratory
tract (44). Traditionally, IgE loading of tissue mast cells has been
thought to occur through a passive diffusion model. If so, we
would have expected mast cells to play some role in IgE clear-
ance. However, mast cell-deﬁcient mice showed overlapping peak
IgE levels and rates of clearance as compared with wild-type mice
(data not shown). Therefore, while similar logic regarding a lim-
ited number of unoccupied receptors is certainly possible, we
speculate that tissue mast cells may have limited access to serum
IgE by virtue of the endothelial barrier. If such limitations in
access existed, mast cells might then be expected to contribute
little to serum IgE clearance, unless local changes in delivery
or vasopermeability occurred. These possibilities require further
The absence of B cells (or blockade of CD23) led to an ∼2-fold
increase in the peak serum IgE concentration (Figs. 3B,4A).
Despite this increase in peak serum IgE, we observed no clear
change in the overall rate of IgE clearance from the serum (Fig.
3B). This lack of change in IgE clearance is in agreement with
published ﬁndings in CD23
mice (45). However, in these
CD23-deﬁcient mice, no difference in peak IgE was observed after
i.v. infusion. This likely reﬂects the 30- to 40-fold larger amount
of infused IgE (∼75–100 mg). The peak IgE level after such
infusions is in the 30–40 mg range, which may exceed physiologic
limits that might have been detected using the amounts we used
(13). Additionally, the use of the epitope-tagged IgE molecule
allowed us to isolate the kinetics of an individual group of IgE
molecules independent of the native IgE pool.
The similar rate of IgE clearance in B cell-deﬁcient mice sug-
gests that IgE homeostasis is regulated on several levels. While
B cells limit the overall size of the IgE pool by rapidly binding free
IgE molecules, other mechanisms exist to determine the catabolism
and clearance of IgE. IgE-binding factors have long been postu-
lated to contribute to the clearance of IgE (8, 21), but a detailed
understanding is lacking. The regulation of IgE levels by CD23
has focused primarily on alterations in IgE production. In partic-
ular, the ﬁrst published CD23-deﬁcient mouse had a moderate 2-
fold increase in total serum IgE, along with an up to 10-fold in-
crease in Ag-speciﬁc IgE production following immunization
(37). Subsequent mouse strains had a mixture of results, including
normal baseline serum IgE levels, no alteration in Ag-speciﬁc IgE
production, and defects in Ag focusing following hapten treatment
in mice presensitized with Ag-speciﬁc IgE (45, 46). These data
supplemented previous work suggesting defective IgE production
in anti-CD23–treated rodents (47). Many of these differences
likely relate to the interstrain variation, genetic backgrounds, and
different experimental model systems, as well as to the inherent
complexity of CD23 biology, including the binding characteristics
of the IgE-CD23 interaction and subsequent effects on B cell
survival (8). In humans, the biology is further complicated by
interactions between CD23 and CD21, which enhance IgE pro-
duction (21, 48, 49).
Anti-CD23 has been considered as a biotherapeutic in human
allergic disease. The impact of anti-CD23 treatment on allergic
disease remains unclear, but early data pointed to a rapid drop in
IgE levels following anti-CD23 treatment (50, 51). Although these
data would appear to counter ours, the Ab targets as well as dif-
ferences in human and mouse CD23 biology, including an ∼10-
fold lower afﬁnity of human CD23 for IgE, may account for these
outcomes (13, 17, 52).
CD23 exists in both a membrane-bound and soluble form. Our
data indicate that B cells can directly bind to serum IgE, although
the low afﬁnity of the interaction required us to perform ﬁxation to
detect this binding. Although our data do not exclude a role for
soluble CD23 in the regulation of IgE levels, previous work with
isolated overexpression of each form of CD23 has suggested that the
membrane-bound form has a much greater impact on IgE biology
than does the soluble form (53). The lower afﬁnity of soluble CD23
for IgE, and an inability of mouse CD23 to interact with CD21,
further diminishes the likelihood of a signiﬁcant role for soluble
CD23 in regulating serum IgE levels or IgE production (49).
The amount of CD23-associated IgE on B cells was directly
proportional to the serum IgE level and showed rapid turnover
(Fig. 5B). Although it is possible that ADAM10-mediated CD23
shedding increases with ligand binding (54, 55), no data yet exist
to demonstrate such a correlation, and ligand binding may in fact
decrease CD23 shedding (13). As our data indicate, IgE molecules
are internalized and likely degraded. This receptor-mediated en-
docytosis has also been demonstrated by in vitro experiments using
murine B cell hybridomas expressing CD23, EBV-transformed hu-
man B cells, and additional cell lines (38, 56, 57).
The size of the free serum IgE pool is a function of production,
clearance of IgE molecules, and capture of IgE by peripheral
cells. Our data indicate that the latter appears to make little
contribution to the regulation of the free IgE pool and that B cells,
through a low-afﬁnity receptor, are a primary regulator of the IgE
pool. Binding of monomeric IgE by B cells likely plays at least
two important roles in the expression of allergic inﬂammation.
FIGURE 7. CD23 blockade leads to an increase in the serum IgE pool.
Groups of 3 BALB/c mice received 100 mg of either isotype control (N)or
B3B4 (:) Ab on day 0. One, 4, and 7 d later, serum was collected from
individual animals and serum IgE levels were examined by ELISA. Error
bars for each time point represent the SEM. Data are representative of two
independent experiments. pp , 0.01.
The Journal of Immunology 5045
First, CD23 binding of monomeric IgE serves to enhance Ag
presentation and Ag focusing, as has been previously described
(46). Serum IgE levels can be detected within the ﬁrst 5–8 d of
an immune response (58, 59), and these Abs could facilitate Ag
capture and presentation by B cells. Second, B cell capture of
serum IgE levels decreases the availability of IgE molecules for
loading onto mast cells in peripheral tissue. Because loss of this
binding capacity enhances hypersensitivity responses (Fig. 6), we
speculate that B cells act as a checkpoint to guard against the
potentially fatal consequences of inappropriate mast cell activa-
tion by IgE and Ag. This system biases mast cells toward loading
with IgE speciﬁcities that are highly represented in the serum.
Presumably, these speciﬁcities would be the most relevant for
host defense, although in the modern age, allergic manifestations
are the more common result. Further work in humans to examine
whether human CD23 plays a similar regulatory role as murine
CD23 and to clarify the precise role of speciﬁc domains of CD23
as well as the cell types involved will give us a better perspective
on the role of CD23 in allergic inﬂammation and insights to target
CD23 function in disease.
We thank J. Shin, G. Caughey, and M. Wabl for mice and reagents and
N. Flores for assistance with animals. We also thank members of the Locks-
ley Laboratory, C. Lowell, A. DeFranco, and C. Allen for thoughtful com-
ments on the manuscript.
The authors have no ﬁnancial conﬂicts of interest.
1. Locksley, R. M. 2010. Asthma and allergic inﬂammation. Cell 140: 777–783.
2. Tsujimura, Y., K. Obata, K. Mukai, H. Shindou, M. Yoshida, H. Nishikado,
Y. Kawano, Y. Minegishi, T. Shimizu, and H. Karasuyama. 2008. Basophils play
a pivotal role in immunoglobulin-G-mediated but not immunoglobulin-E-
mediated systemic anaphylaxis. Immunity 28: 581–589.
3. Miyajima, I., D. Dombrowicz, T. R. Martin, J. V. Ravetch, J. P. Kinet, and
S. J. Galli. 1997. Systemic anaphylaxis in the mouse can be mediated largely
through IgG1 and FcgRIII: assessment of the cardiopulmonary changes,
mast cell degranulation, and death associated with active or IgE- or IgG1-
dependent passive anaphylaxis. J. Clin. Invest. 99: 901–914.
4. Dombrowicz, D., V. Flamand, I. Miyajima, J. V. Ravetch, S. J. Galli, and
J. P. Kinet. 1997. Absence of FcεRI a chain results in upregulation of FcgRIII-
dependent mast cell degranulation and anaphylaxis: evidence of competition
between FcεRI and FcgRIII for limiting amounts of FcR b and g chains. J. Clin.
Invest. 99: 915–925.
5. Stone, S. P., G. J. Gleich, and S. A. Muller. 1976. Atopic dermatitis and IgE:
relationship between changes in IgE levels and severity of disease. Arch. Der-
matol. 112: 1254–1255.
thrich, B. 1978. Serum IgE in atopic dermatitis: relationship to severity of
cutaneous involvement and course of disease as well as coexistence of atopic
respiratory diseases. Clin. Alle rgy 8: 241–248.
7. Abramson, J., and I. Pecht. 2007. Regulation of the mast cell response to the type
1Fcε receptor. Immunol. Rev. 217: 231–254.
8. Corry, D. B., and F. Kheradmand. 1999. Induction and regulation of the IgE
response. Nature 402(6760 Suppl.)B18–B23.
9. Gould, H. J., and B. J. Sutton. 2008. IgE in allergy and asthma today. Nat. Rev.
Immunol. 8: 205–217.
10. Kitaura, J., T. Kinoshita, M. Matsumoto, S. Chung, Y. Kawakami, M. Leitges,
D. Wu, C. A. Lowell, and T. Kawakami. 2005. IgE- and IgE+Ag-mediated
mast cell migration in an autocrine/paracrine fashion. Blood 105: 3222–3229.
11. Kraft, S., and J. P. Kinet. 2007. New developments in FcεRI regulation, function
and inhibition. Nat. Rev. Immunol. 7: 365–378.
12. Kawakami, T., and S. J. Galli. 2002. Regulation of mast-cell and basophil
function and survival by IgE. Nat. Rev. Immunol. 2: 773–786.
13. Conrad, D. H., J. W. Ford, J. L. Sturgill, and D. R. Gibb. 2007. CD23: an
overlooked regulator of allergic disease. Curr. Allergy Asthma Rep. 7: 331–337.
14. Munoz, O., C. Brignone, N. Grenier-Brossette, J. Y. Bonnefoy, and J. L. Cousin.
1998. Binding of anti-CD23 monoclonal antibody to the leucine zipper motif of
FcεRII/CD23 on B cell membrane promotes its proteolytic cleavage: evidence
for an effect on the oligomer/monomer equilibrium. J. Biol. Chem. 273: 31795–
15. Maeda, K., G. F. Burton, D. A. Padgett, D. H. Conrad, T. F. Huff, A. Masuda,
A. K. Szakal, and J. G. Tew. 1992. Murine follicular dendritic cells and low
afﬁnity Fc receptors for IgE (FcεRII). J. Immunol. 148: 2340–2347.
16. LeBien, T. W., and T. F. Tedder. 2008. B lymphocytes: how they develop and
function. Blood 112: 1570–1580.
17. Zhang, M., R. F. Murphy, and D. K. Agrawal. 2007. Decoding IgE Fc receptors.
Immunol. Res. 37: 1–16.
18. Hjelm, F., M. C. Karlsson, and B. Heyman. 2008. A novel B cell-mediated
transport of IgE-immune complexes to the follicle of the spleen. J. Immunol.
19. Yu, L. C., P. C. Yang, M. C. Berin, V. Di Leo, D. H. Conrad, D. M. McKay,
A. R. Satoskar, and M. H. Perdue. 2001. Enhanced transepithelial antigen
transport in intestine of allergic mice is mediated by IgE/CD23 and regulated by
interleukin-4. Gastroenterology 121: 370–381.
20. Bonnefoy, J. Y., S. Lecoanet-Henchoz, J. F. Gauchat, P. Graber, J. P. Aubry,
P. Jeannin, and C. Plater-Zyberk. 1997. Structure and functions of CD23. Int.
Rev. Immunol. 16: 113–128.
21. Hibbert, R. G., P. Teriete, G. J. Grundy, R. L. Beavil, R. Reljic, V. M. Holers,
J. P. Hannan, B. J. Sutton, H. J. Gould, and J. M. McDonnell. 2005. The structure of
human CD23 and its interactions with IgE and CD21. J. Exp. Med. 202: 751–760.
22. Vieira, P., and K. Rajewsky. 1988. The half-lives of serum immunoglobulins in
adult mice. Eur. J. Immunol. 18: 313–316.
23. Achatz-Straussberger, G., N. Zaborsky, S. Ko
nigsberger, S. Feichtner, S. Lenz,
D. Peckl-Schmid, M. Lamers, and G. Achatz. 2009. Limited humoral immu-
noglobulin E memory inﬂuences serum immunoglobulin E levels in blood. Clin.
Exp. Allergy 39: 1307–1313.
24. Dreskin, S. C., P. K. Goldsmith, W. Strober, L. A. Zech, and J. I. Gallin. 1987.
Metabolism of immunoglobulin E in patients with markedly elevated serum
immunoglobulin E levels. J. Clin. Invest. 79: 1764–1772.
25. Iio, A., T. A. Waldmann, and W. Strober. 1978. Metabolic study of human IgE:
evidence for an extravascular catabolic pathway. J. Immunol. 120: 1696–1701.
26. Dombrowicz, D., V. Flamand, K. K. Brigman, B. H. Koller, and J. P. Kinet. 1993.
Abolition of anaphylaxis by targeted disruption of the high afﬁnity immuno-
globulin E receptor a chain gene. Cell 75: 969–976.
27. Mohrs, M., K. Shinkai, K. Mohrs, and R. M. Locksley. 2001. Analysis of type 2
immunity in vivo with a bicistronic IL-4 reporter. Immunity 15: 303–311.
28. Kaplan, M. H., U. Schindler, S. T. Smiley, and M. J. Grusby. 1996. Stat6 is
required for mediating responses to IL-4 and for development of Th2 cells.
Immunity 4: 313–319.
29. McKenzie, G. J., P. G. Fallon, C. L. Emson, R. K. Grencis, and A. N. McKenzie.
1999. Simultaneous disruption of interleukin (IL)-4 and IL-13 deﬁnes individual
roles in T helper cell type 2-mediated responses. J. Exp. Med. 189: 1565–1572.
30. Wolters, P. J., J. Mallen-St Clair, C. C. Lewis, S. A. Villalta, P. Baluk, D. J. Erle,
and G. H. Caughey. 2005. Tissue-selective mast cell reconstitution and differ-
ential lung gene expression in mast cell-deﬁcient Kit
sash mice. Clin.
Exp. Allergy 35: 82–88.
31. Mukai, K., K. Matsuoka, C. Taya, H. Suzuki, H. Yokozeki, K. Nishioka,
K. Hirokawa, M. Etori, M. Yamashita, T. Kubota, et al. 2005. Basophils play
a critical role in the development of IgE-mediated chronic allergic inﬂammation
independently of T cells and mast cells. Immunity 23: 191–202.
32. Mathur, A., D. H. Conrad, and R. G. Lynch. 1988. Characterization of the murine
T cell receptor for IgE (FcεRII): demonstration of shared and unshared epitopes
with the B cell FcεRII. J. Immunol. 141: 2661–2667.
33. Cernadas, M., G. T. De Sanctis, S. J. Krinzman, D. A. Mark, C. E. Donovan,
J. A. Listman, L. Kobzik, H. Kikutani, D. C. Christiani, D. L. Perkins, and
P. W. Finn. 1999. CD23 and allergic pulmonary inﬂammation : potential role as
an inhibitor. Am. J. Respir. Cell Mol. Biol. 20: 1–8.
34. Wan, T., R. L. Beavil, S. M. Fabiane, A. J. Beavil, M. K. Sohi, M. Keown,
R. J. Young, A. J. Henry, R. J. Owens, H. J. Gould, and B. J. Sutton. 2002. The
crystal structure of IgE Fc reveals an asymmetrically bent conformation. Nat.
Immunol. 3: 681–686.
35. Zhu, D., C. L. Kepley, M. Zhang, K. Zhang, and A. Saxon. 2002. A novel human
immunoglobulin Fcg-Fcε bifunctional fusion protein inhibits FcεRI-mediated
degranulation. Nat. Med. 8: 518–521.
36. Gessner, A., K. Mohrs, and M. Mohrs. 2005. Mast cells, basophils, and eosi-
nophils acquire constitutive IL-4 and IL-13 transc ripts during lineage differen-
tiation that are sufﬁcient for rapid cytokine production. J. Immunol. 174: 1063–
37. Yu, P., M. Kosco-Vilbois, M. Richards, G. Ko
hler, and M. C. Lamers. 1994.
Negative feedback regulation of IgE synthesis by murine CD23. Nature 369:
38. Chen, S. S. 1991. Mechanisms of IgE homeostasis: sequestra tion of IgE by
murine type II IgE Fc receptor-bearing B cell hybridomas. J. Immunol. 147:
39. Zaidi, A. K., and D. W. MacGlashan. 2010. Regulation of FcεRI expression
during murine basophil maturation: the interplay between IgE, cell division, and
FcεRI synthetic rate. J. Immunol. 184: 1463–1474.
40. Kubo, S., K. Matsuoka, C. Taya, F. Kitamura, T. Takai, H. Yonekawa, and
H. Karasuyama. 2001. Drastic up-regulation of Fcepsilonri on mast cells is in-
duced by IgE binding through stabilization and accumulation of FcεRI on
the cell surface. J. Immunol. 167: 3427–3434.
41. Sokol, C. L., N. Q. Chu, S. Yu, S. A. Nish, T. M. Laufer, and R. Medzhitov. 2009.
Basophils function as antigen-presenting cells for an allergen-induced T helper
type 2 response. Nat. Immunol. 10: 713–720.
42. Sullivan, B. M., and R. M. Locksley. 2009. Basophils: a nonredundant con-
tributor to host immunity. Immunity 30: 12–20.
43. Sokol, C. L., and R. Medzhitov. 2010. Role of basophils in the initiation of Th2
responses. Curr. Opin. Immunol. 22: 73–77.
5046 B CELL REGULATION OF SERUM IgE
44. Galli, S. J., and M. Tsai. 2010. Mast cells in allergy and infection: versatile
effector and regulatory cells in innate and adaptive immunity. Eur. J. Immunol.
45. Stief, A., G. Texido, G. Sansig, H. Eibel, G. Le Gros, and H. van der Putten.
1994. Mice deﬁcient in CD23 reveal its modulatory role in IgE production but no
role in T and B cell development. J. Immunol. 152: 3378–3390.
46. Fujiwara, H., H. Kikutani, S. Suematsu, T. Naka, K. Yoshida, K. Yoshida,
T. Tanaka, M. Suemura, N. Matsumoto, and S. Kojima. 1994. The absence of IgE
antibody-mediated augmentation of immune responses in CD23-deﬁcient mice.
Proc. Natl. Acad. Sci. USA 91: 6835–6839.
47. Flores-Romo, L., J. Shields, Y. Humbert, P. Graber, J. P. Aubry, J. F. Gauchat,
G. Ayala, B. Allet, M. Chavez, H. Bazin, et al. 1993. Inhibition of an in vivo
antigen-speciﬁc IgE response by antibodies to CD23. Science 261: 1038–1041.
48. Aubry, J. P., S. Pochon, P. Graber, K. U. Jansen, and J. Y. Bonnefoy. 1992. CD21
is a ligand for CD23 and regulates IgE production. Nature 358: 505–507.
, R., G. Cosentino, and H. J. Gould. 1997. Function of CD23 in the re-
sponse of human B cells to antigen. Eur. J. Immunol. 27: 572–575.
50. Rosenwasser , L. J., W . W. Busse, R. G. Lizambri, T . A. Olejnik, and M. C. Totoritis.
2003. Allergic asthma and an anti-CD23 mAb (IDEC-152): results of a phase I,
single-dose, dose-escalating clinical trial. J. Allergy Clin. Immunol. 112: 563–570.
51. Rosenwasser, L. J., and J. Meng. 2005. Anti-CD23. Clin. Rev. Allergy Immunol.
52. Conrad, D. H. 1990. FcεRII/CD23: the low afﬁnity receptor for IgE. Annu. Rev.
Immunol. 8: 623–645.
53. Texido, G., H. Eibel, G. Le Gros, and H. van der Putten. 1994. Transgene CD23
expression on lymphoid cells modulates IgE and IgG1 responses. J. Immunol.
54. Lemieux, G. A., F. Blumenkron, N. Yeung, P. Zhou, J. Williams, A. C. Grammer,
R. Petrovich, P. E. Lipsky, M. L. Moss, and Z. Werb. 2007. The low afﬁnity IgE
receptor (CD23) is cleaved by the metalloproteinase ADAM10. J. Biol. Chem.
55. Weskamp, G., J. W. Ford, J. Sturgill, S. Martin, A. J. Docherty, S. Swendeman,
N. Broadway, D. Hartmann, P. Saftig, S. Umland, et al. 2006. ADAM10 is a
principal “sheddase” of the low-afﬁnity imm unoglobulin E receptor CD23. Nat.
Immunol. 7: 1293–1298.
56. Karagiannis, S. N., J. K. Warrack, K. H. Jennings, P. R. Murdock, G. Christie,
K. Moulder, B. J. Sutton, and H. J. Gould. 2001. Endocytosis and recycling of the
complex between CD23 and HLA-DR in human B cells. Immunology 103: 319–331.
57. Montagnac, G., A. Molla
-Herman, J. Bouchet, L. C. Yu, D. H. Conrad,
M. H. Perdue, and A. Benmerah. 2005. Intracellular trafﬁcking of CD23: dif-
ferential regulation in humans and mice by both extracellular and intracellular
exons. J. Immunol. 174: 5562–5572.
58. Morris, S. C., R. L. Coffman, and F. D. Finkelman. 1998. In vivo IL-4 responses
to anti-IgD antibody are MHC class II dependent and b
dependent and develop normally in the absence of IL-4 priming of T cells. J.
Immunol. 160: 3299–3304.
59. Thyphronitis, G., I. M. Katona, W. C. Gause, and F. D. Finkelman. 1993.
Germline and productive C epsilon gene expression during in vivo IgE
responses. J. Immunol. 151: 4128–41 36.
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