Proc. NatL Acad. Sci. USAVol. 80, pp. 293-297, January 1983Medical Sciences

Induction of a B-lymphocyte receptor for aT cell-replacing factorby the crosslinking of surface IgD

(B-cell differentiation/T-cell helper factors/anti-immunoglobulin antibody/immune-defective B cells/cell factor. adsorptions)

L. J. YAFFE AND FRED D. FINKELMANPathology Branch, Naval Medical Research Institute, and the Department of Medicine, Uniformed Services University of the Health Sciences,Bethesda, Maryland 20814

Communicated by William E. Paul, October 4, 1982

ABSTRACT The observation that anti-immunoglobulin anti-bodies and T cell-replacing factor (TRF) have a synergistic effecton the stimulation ofB lymphocytes to differentiate into antibody-secreting cells suggested to us the possibility that the crosslinkingof B-cell surface immunoglobulin by antigen or anti-immunoglob-ulin antibody might induce the expression of a B-cell receptor forTRF. In order to test this possibility we studiedwhether spleencells from mice injected with 400-800 ,ug of anti-IgD antibody 1-3 days before sacrifice had an enhanced capacity to adsorb TRFactivity from a partially purified culture supernatant of conca-navalin A-stimulated mouse spleen cells. We found that spleencells from euthymic or congenitally athymic mice injected 24-30hr prior to sacrifice with either affinity-purified goat anti-mouseIgD antibody or a monoclonal allospecific anti-IgD antibody had> 100 times the TRF adsorptive capacity of spleen cells from con-trol mice. In contrast, spleen cells from anti-IgD treated DBA/2Ha mice, which have been shown to have an immune defect as-sociated with the lack of a TRF receptor, were unable to adsorbTRF activity from concanavalin A-stimulated helper superna-tants. This suggests that the crosslinking of B-cell surface immu-noglobulin may induce B lymphocytes to express a receptor orreceptors for TRF and thus enhance B-cell responsiveness to thishelper factor.

The crosslinking ofB-lymphocyte surface immunoglobulin. (sIg)by antigen or anti-Ig antibody has a number of effects that con-tribute to a humoral immune response. These effects includethe induction of an increase in B-cell proliferation (1-5), a 2- to3-fold increase in the expression of B-cell surface Ia antigen (6),and enhancement of the ability of B cells to present antigensto T cells (7, 8). None of these effects, however, fully explainsthe greatly increased antibody response seen when B lympho-cytes are cultured with anti-Ig antibody plus a supernatant fromconcanavalin A (Con A)-stimulated T lymphocytes that containsT cell-replacing factor (TRF) activity, as opposed to culture withthis TRF-containing supernatant in the absence of anti-Ig an-tibody (9, 10).One possible explanation for the synergistic effects of anti-Ig

antibody and Con A-induced TRF on the differentiation of Blymphocytes into antibody-secreting cells is that sIg crosslink-ing by antigen or anti-Ig might induce not only B-cell prolif-eration but also the expression of B-cell receptors for TRF andthus facilitate the action of TRF on B lymphocytes. Becauseinjection of mice with an antibody to IgD has a number of poly-clonal activating effects on B lymphocytes (11, 12), we decidedto study the ability of anti-IgD to induce B-cell receptors forTRF, as measured by the capacity of spleen cells from anti-IgDor control Ig injected mice to adsorb TRF activity from the su-pernatant of Con A-stimulated spleen cells.

We have found that spleen cells from normal or congenitallyathymic mice that had been injected with an allospecific mono-clonal or a heterologous affinity-purified anti-6 antibody 24 hrbefore sacrifice adsorb TRF with more than 100-fold greaterability than control spleen cells. Spleen cells from mice with theDBA/2Ha immune defect, which have been shown to lack areceptor for TRF (13, 14), were unable to adsorb TRF activityeven after activation by in vivo treatment with anti-IgDlanti-body. We conclude that one effect of ligand crosslinking of sIgon normal B-lymphocyte sIg may be the induction ofTRF re-ceptor expression.

MATERIAL AND METHODSMice. DBA/2, BALB/c, and (BALB/c X C57BL/6)Fl

(BCF1) mice were purchased from The Jackson Laboratory.Swiss nu/nu mice were purchased from the Small Animal Sec-.tion of the National Institutes of Health (Bethesda, MD).(DBA/2Ha X DBA/2)F1 male and female mice were originallyobtained from West Seneca Laboratories (Buffalo; NY), andwere bred and maintained at Dominion Laboratories (Dublin,VA).

T-Cell Depletion.. Spleen cell suspensions were T-cell de-pleted by a double treatment with monoclonal anti-Thy 1.2 an-tibody (kindly supplied by Philip.Lake) (15) together with a 1:15dilution of rabbit complement that had been adsorbed withmouse spleen and thymus cells. Cell sorter analysis showed that<1% of the cells remaining after this treatment were Thy 1.2+.Furthermore, these cells gave no proliferative response to ConA as measured. by in vitro thymidine incorporation.

Cell Cultures. One million T cell-depleted spleen cells from8- to 12-week-old mice were -cultured in 0.2 ml of modifiedMishell-Dutton medium (16) containing 10% fetal bovineserum and 50 ,kM 2-mercaptoethanol in flat-bottom microtiterplate wells (Costar, Cambridge; MA). Cultures were incubatedstationary at 37°C for 4 days in a humidified 5% CO2/95% airatmosphere. Adsorbed and unadsorbed TRF-containing super-natants were added to cultures on day. 2.

Antigens. Sheep. erythrocytes (SRBC) were obtained from asingle-animal. One million SRBC were routinely used per cellculture.

Plaque-Forming Cell (pfc) Assays. Cultured cells were har-vested from microtiter plate wells after 4 days ofincubation andthe number of specific IgM pfc was determined by the tech-nique of Cunningham and. Szenberg (17). Data represent geo-metric means x/-. SEM of triplicate cultures. Backgroundplaques with no antigen were less than 6 per 106 cultured cellsin the presence or absence of adsorbed or unadsorbed TRF.

Abbreviations: sIg, surface immunoglobulin; Con A, concanavalin A;TRF, T cell-replacing factor; SRBC, sheep erythrocytes; pfc, plaque-forming cell(s).

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Anti-IgD Antibody. Goat anti-mouse 8 was prepared andaffinity-purified as described (18). A monoclonal IgG2b im-munoglobulin (H8a/1) with specificity for mouse IgD of the a

but not ofthe b allotype was purified from ascitic fluid ofC. B20mice harboring this hybridoma (a generous gift of Ian Zitron)by adsorption to and elution from protein A-Sepharose (19).Normal goat IgG, affinity-purified goat anti-ferritin, or CBPC-101, a monoclonal IgG2a of the b allotype that does not reactwith mouse cells, were used as control antibodies (11).

Preparation of TRF. TRF-containing Con A supernatantswere prepared essentially as described by Schimpl and Wecker(20). DBA/2 or BALB/c spleen cells (106/ml) were culturedin RPMI-1640 supplemented with 20 mM Hepes, 2 mM L-glu-tamine, 0.075% NaHCO3, 25 pLM 2-mercaptoethanol, genta-micin at 0.05 mg/ml and Con A at 1 ,ug/ml. Cultures -were

maintained for 48 hr in 75-cm2 culture flasks (Costar) in a 7.5%C02/92.5% air humidified atmosphere at 37C. Culture su-

pernatants were concentrated by pressure ultrafiltration throughan Amicon PM-10 membrane and were fractionated by Seph-adex G-100 gel filtration. The 40,000 ± 7,500 apparent molec-ular weight fraction was collected and designated "TRF. ' Theseprocedures removed essentially 99% ofCon A from the super-

natant as indicated by the removal ofa radiolabeled Con A tracerduring the procedure. Although the supernatant was deter-mined to have virtually no interleukin 2 activity on the basis ofa proliferation assay using an interleukin 2-dependent cell line(data not shown) (21), the supernatant was highly active at re-

constituting in vitro IgM anti-SRBC antibody responses of Tcell-depleted spleen cells when added on day 2 of culture. Forthis reason, and for the sake of simplicity, we are calling thissupernatant TRF even though it quite likely contains more thanone active lymphokine.

Adsorption of TRF Reconstituting Activity. By proceduressimilar to those used by Tominaga and co-workers (13, 14), TRFwas adsorbed by spleen cells from anti-IgD-treated or controlmice. Spleen cells were prepared from DBA/2 mice injectedintravenously 1-3 days prior with 800 Ag ofgoat anti-mouse IgDor 400 ug of monoclonal anti-IgD or equal quantities of the ap-

propriate control antibody. Spleen cells (1 x 1i0 to 1 X 108)were incubated at 370C for 30 min with 1 ml ofTRF, followedby centrifugation at 1,200 x g for 10 min to remove cells. Asecond adsorption was then performed with an equal numberof fresh spleen cells from anti-IgD or control antibody-treatedmice. Supernatants were sterilized by filtration and stored at-700C. Culture media adsorbed as above with spleen cells from

anti-IgD or control antibody treated mice had no inhibitory-orstimulatory effect on the ability of unadsorbed TRF to recon-

stitute T cell-depleted spleen cell responses.

RESULTSIn order to determine whether anti-1gD treatment inducedspleen cells to acquire a receptor for one or more helper factorspresent in the partially purified supernatant of Con A-stimu-lated spleen cells, aliquots of this supernatant were adsorbedtwice with 105, 106, 107, or 10' spleen cells from DBA/2 miceinjected 1 or 3 days earlier with'800 pLg of goat anti-mouse IgDor control antiserum and then assayed for their late-acting TRFactivity. This was measured by their ability to reconstitute an

in vitro antibody response to SRBC for T cell-depleted spleencells when added 2 days after the start ofculture. Con A-inducedhelper supernatants adsorbed twice with 105 to 10' spleen cellsfrom mice injected 1-3 days earlier with control antibody didnot differ inTRF activity (Table 1). In contrast, the TRF activityof supernatants adsorbed twice with 106 or more spleen cellsfrom mice injected 1-3 days earlier with 800 jig of goat anti-mouse IgD was reduced by 60-82%. Two additional experi-ments (data not shown) gave similar results.

In order to evaluate the possibility that supernatants ad-sorbed with spleen cells from anti-IgD treated mice might con-

tain factors that suppressed the generation ofantibody-secretingcells, the effect of adding 20 pl of supernatant adsorbed with108 spleen cells to 5-100 p1 of unadsorbed TRF was examined.These experiments showed that the adsorbed supernatant hadno suppressive effect at any concentration ofunadsorbed helpersupernatant used (data not shown). Thus, spleen cells from mice

injected 1 day prior to sacrifice have >100 times the capacityto adsorbTRF from a Con A-stimulated helper supernatant thando control spleen cells. Additional experiments in which thehelper activities of varying quantities of adsorbed and unad-sorbed Con A supernatants were compared indicated that thereduction in TRF activity obtained by adsorption of a Con Ahelper supernatant with 106 spleen cells from day 1 anti-IgD-treated mice reduced TRF activity by approximately 80% (datanot shown).

In order to establish whether the increase in TRF-adsorbingcapacity induced by the injection of goat anti-mouse IgD re-

sulted from the binding and crosslinking ofB-cell sIgD, BALB/c, C57BL/6, or BCF, mice were injected with H8a/l, a mono-

clonal mouse IgG2b of the b allotype that binds. to IgD of thea allotype (found in BALB/c mice) but has little or no bindingcapacity for IgD ofthe b allotype (found in C57BL/6 mice) (19).Spleen cells from BALB/c mice injected 24 hr prior to sacrificewith 400 pzg of H8a/1 anti-6 had more than 100 times the TRF-adsorbing capacity of BALB/c spleen cells from mice. injectedwith a control monoclonal antibody (Table 2). On the otherhand, only minor differences were seen in the TRF-adsorbingcapacity of spleen cells from C57BL/6 mice injected with H8a/

Table 1. Adsorption of TRF activity with spleen cells from mice treated with goat anti-mouse IgD

TRF adsorption with Anti-SRBC pfc/106 cultured T cell-depleted DBA/2 spleen cells,DBA/2 spleen cells with different numbers of spleen cells from treated mice

Days after Treated used for TRF adsorptiontreatment with 1 x 105 1xx106 1 x lo8

1 NGIg 59 (1.22) 50 (1.26) 58 (1.23) 54 (1.12)GaMS 43 (1.13) 20 (1.15) 13 (1.36) 10 (1.19)

3 NGIg 53 (1.35) 65 (1.19) 59 (1.11) 61 (1.23)GaMS 68 (1.09) 19 (1.27) 15 (1.19) 13 (1.18)

Spleen cells used for TRF adsorption were from mice treated 1 or 3 days earlier with 800 ug of goatanti-mouse IgD (GaM8) or 800 ,.g of normal goat Ig (NGIg). Spleen cells (1 x 10r to 1 x 108) from treatedand control mice were used to adsorb 1 ml of TRF supernatant, and residual TRF activity was assayedon T cell-depleted spleen cells. TRF was added on day 2 of 4-day cultures for optimal pfc responses, withTRF representing 25% of culture volume. Data represent geometric means (with x/+ SEM given inparentheses) for triplicate cultures with background plaques (<3 pfc) subtracted.

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Table 2. Adsorption of TRF activity with spleen cells from mice treated 24 hr earlier withmonoclonal H6/1 anti-IgD

Anti-SRBC pfc/106 cultured T cell-depleted DBA/2spleen cells, with different numbers of spleen cells

TRF adsorption with from treated mice used for TRF adsorptionspleen cells from Treated with 1 x 105 1 x 106 1 x 107 1 x 108

BALB/c CBPC-101 99 (1.09) 102 (1.14) 102 (1.06) 96 (1.13)H5/1 93 (1.04) 50 (1.11) 41 (1.12) 32 (1.15)

C57BL/6 CBPC-101 96 (1.17) 89 (1.14) 94 (1.11) 89 (1.08)HSa/1 93 (1.16) 80 (1.15) 82 (1.08) 68 (1.04)

BCF1 CBPC-101 99 (1.11) 85 (1.15) 94 (1.08) 95 (1.12)H8/1 90 (1.08) 68 (1.14) 55 (1.18) 46 (1.09)

Mice were treated with 400 yg of monoclonal H54/1 anti-IgD or CBPC-101. TRF was added on day 2of 4-day cultures for optimal pfc responses, with TRF representing 25% of culture volume. Data are pre-sented as in Table 1; background plaques (<5 pfc) are subtracted.

1 anti-S or with control antibody. The spleen cells of BCF1 mice,whose sIgD' cells are equally divided between the a and b al-lotypes (22), clearly had their ability to adsorb TRF enhancedby in vivo exposure -to H6'/1 anti-S, although the degree ofenhancement was less than that seen with BALB/c mice. Be-cause Haa/l anti-S binds to the sIgD on BALB/c and BCF1 Bcells but not toC57BL/6 B-cell slgD, these studies indicate thatthe TRF receptor-enhancing activity of anti-IgD antibody re-sults from its interaction with B-cell sIgD.The incomplete adsorption ofTRF activity by even 108 H8'/

1 anti-S treated BALB/c spleen cells in this experiment mayindicate that more than one lymphokine was responsible for theTRF activity in the helper supernatant used and that anti-IgDstimulated receptor expression for only one of these lympho-kines. Alternatively, a very small number of T cells that mayhave renmained viable in the cultured cell population after tworounds of killing with anti-Thy 1.2' and complement may havegenerated TRF in response to another lymphokine in the ConA-stimulated helper supernatant that did not bind to anti-IgDstimulated spleen cells.

In additional experiments, we investigated the T-cell depen-dence of the anti-IgD-induced enhancement of TRF-adsorbingcapacity by studing the effect of goat anti-mouse IgD on theability of spleen cells from congenitally athymic (nu/nu) miceto adsorb TRF. Spleen cells from nu/nu mice that were injected30 hr before sacrifice with goat anti-mouse IgD had >100 timesmore TRF-adsorbing capacity than spleen cells from nu/numice injected with a control antibody (Table 3). Two additionalexperiments gave similar results. Thus, the enhancing effect ofanti-IgD treatment on the adsorption ofTRF by spleen cells isT-cell independent, at least to the extent that nu/nu mice aredevoid ofT lymphocytes.

Table 3. Adsorption of TRF activity with spleen cells from Swissnu/nu mice treated 30 hr earlier with goat anti-mouse IgD

TRF adsorption Anti-SRBC pfc/106 cultured T cell-depletedwith DBA/2 spleen cells, with different numbers

Swiss nu/nu of spleen cells from treated mice usedspleen cells for TRF adsorptiontreated with lx 105 1 x 106 1 x 107 1 x 108

GaF 48 (1.16) 59 (1.14) 63 (1.23) 70 (1.12)GaMs 35 (1.19) 3 (2.04) 4 (2.12) 1(1.00)

Spleen cells for TRF adsorption were made from nude mice treated30 hr earlier with 800 ,ug of goat anti-mouse IgD (GaM8) or 800 Mgof antiferritin (GaF) antibody. TRF was added on day 2 of 4-day cul-tures for optimal pfc responses, with TRF representing 25% of culturevolume. Data are presented as in Table 1; background plaques (<6 pfc)are subtracted.

In order to determine whether the removal ofTRF activityfrom a Con A supernatant by spleen cells from anti-IgD-treatedmice resulted from the generation ofa receptor specific for TRF,we examined the abilities of spleen cells from anti-IgD-treated(DBA/2Ha x DBA/2)F1 male and female mice to adsorb TRFactivity. This was done because B lymphocytes from DBA/2Ha mice have an X-linked recessive defect associated with theabsence of a TRF-specific receptor and neither respond to noradsorb purified TRF but are normal in many other respects (13,14). Thus, a TRF receptor should be inducible on B cells from(DBA/2Ha x DBA/2)F1 female mice (heterozygous for the im-mune defect) but not on B cells from F1 male mice (hemizygousfor the immune defect). Indeed, spleen cells from the anti-IgD-treated F1 female mice demonstrated a clear ability to adsorbTRF activity from a Con A supernatant but spleen cells fromanti-IgD-treated F1 male mice had no detectable TRF adsorp-tive capacity (Table 4). A second experiment (data not shown)gave similar results. In contrast to this, additional experimentsindicated that, 1 day after injection of anti-IgD, (DBA/2HaX DBA/2)F1 male and female spleen cells showed nearly iden-tical. increases in B-cell expression of surface Ia antigen, spleencell size, and spleen cell DNA synthesis (data not shown).

DISCUSSIONOur studies indicate that the in vivo crosslinking of B-lympho-cyte sIgD by either heterologous or monoclonal homologousanti-3 antibodies induces a >100-fold increase in the ability ofspleen cells from normal mice, but not from mice with the I)BA/2Ha-associated immune defect, to deplete TRF activity froma partially purified supernatant of Con A-stimulated spleencells. This depletion of TRF activity results from the loss ofoneor more lymphokines with this activity rather than the gener-ation of suppressive factors during the adsorption procedurebecause the addition of adsorbed supernatant to unadsorbedsupernatant did not diminish theTRF activity ofthe unadsorbedsupernatant alone. These results strongly suggest that one effectof sIg crosslinking is the induction ofa receptor for one or morelymphokines that have TRF activity. Such an effect would ex-plain the synergistic stimulation ofB-cell differentiation into Ig-secreting cells by anti-Ig antibodies and Con A-induced TRF(9, 10). However, the possibility that the Con A-induced TRFthat replaces T-cell function in an in vitro antibody response toSRBC, as detected in our assay system, differs from the ConA-induced TRF that causes anti-Ig-stimulated B cells to differ-entiate into Ig-secreting cells cannot be eliminated at this time.It should also be noted that some TRFs have been found to haveequal stimulatory effects on antibody production by anti-Ig-treated and untreated B cells (10). It may be that B cells bear

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Table 4. Adsorption of TRF activity with spleen cells from immune.defective (DBA/2Ha xDBA/2)F1 male, and female mice treated 1 day earlier with goat anti-mouse IgD

Anti-SRBC pfc/106 cultured T cell-depleted DBA/2TRF adsorption spleen cells, with different numbers of spleen cellswith spleen from treated mice used for TRF adsorptioncells from Treated with 1 x 105 1 x 106 1 x 107 1 x 108

(DBA/2Ha x DBA/2)F1female GaF 62 (1.10) 69 (1.11) 74 (1.13) 86 (1.15)

GaMS 70 (1.10) 41 (1.15) 11 (1.38) 8 (1.61)(DBA/2Ha x DBA/2)F1male GaF 80 (1.15) 62 (1.11) 67 (1.11) 63 (1.11)

GaMS 75 (1.14) 73 (1.08) 68 (1.17) 70 (1.12)

Spleen cells for TRF adsorption were from mice treated 1 day earlier with 800 pug of goat anti-mouseIgD (GaM8) or 800 ug of goat antiferritin (GaF). Spleen cells (1 x 105 to 1 x 108) from treated and controlmice were used to adsorb 1 ml of TRF supernatant, and residual TRF activity was assayed on T cell-de-pleted DBA/2 spleen cells. TRF was added on day 2 of 4-day cultures for optimal pfc responses, with TRFrepresenting 25% of culture volume. Data are presented as in Table 1; background plaques (<1 pfc) aresubtracted.

both constitutively expressed receptors for some lymphokineswith TRF activity and inducible receptors for other TRFs.

Although we have not yet formally demonstrated that theTRF receptor is present on the B lymphocyte, the finding thatanti-IgD greatly stimulates the. ability of spleen cells from nu/nu mice to adsorb TRF is consistent with this hypothesis. Thefinding that a maximal enhancement of TRF-adsorbing capacityis.seen a relatively short time (24 hr or less) after anti-IgD in-jection is also consistent with the, concept that the increase inTRF adsorptive capacity results directly from a B cell sIg-ligandinteraction rather than from a secondary effect of this interac-tion. We cannot rule out the possibilities, however, that mac-rophages or other non-B, non-T cells play a role in adsorbinga factor thatis required forTRF activity or in inducing anti-IgD-stimulated B lymphocytes to acquire a TRF receptor.

Induction of a receptor or receptors for TRF is only one ofa number of independent mechanisms by which the antigen-induced crosslinking of B-cell sIg can lead to the increased pro-duction of antibody to that antigen. For example, the inductionof B-cell proliferation by the crosslinking of sIg can increase thesize of antigen-reactive B-cell clones (1-5), and the inductionofincreased B-cell surface Ia expression by sIg crosslinking.mayhave a role in enhancing antigen presentation by B cells to T.cells or B-cell receptivity to Ia-restricted helper factors (6-8).We think that the anti-IgD..induced increase in TRF receptorsites of B lymphocytes is independent of the anti-IgD-inducedincrease in B cell surface Ia expression because (i) TRF is notH-2 restricted in its helper activity (23), (ii) in vivo anti-IgD in-jection increases surface Ia expression 2- to 3-fold (6) but in-creases TRF adsorptive capacity >100-fold, and (iii) anti-IgDinduces an increase in the Ia expression but not TRF adsorptive.capacity of spleen cells from mice with the DBA/2Ha immunedefect: Our experimentswith these.immune defective mice alsorule-out the possibility thatTRF is destroyed by incubation withactivated B cells rather than specifically adsorbed because TRFactivity was-noteliminated by adsorption with spleen cells fromanti-IgD-treated (DBA/2Ha X DBA/2)F1 male mice despitethe fact that these cells were as .activated in terms of size andDNA synthesis as spleen cells from anti-IgDl-treated Fi femalemice. Furthermore, the induction ofspleen cellTRF adsorptivecapacity by sIgD crosslinlking is specific to the extent that it doesnot extend to increased-adsorptive capacity for all lymphokines;spleen cells from-mice injected 1 day prior to.,sacrifice with anti-IgD do not have increased ability. to adsorb interleukin.2 (un-published data).

Finally, it should be pointed out that there may be mecha-nisms other than sIg crosslinking that can induce an increasein B-cell TRF receptors. We have recently found that the im-munization of a mouse with SRBC induces its spleen cells tohave an increased TRF adsorptive capacity 7 days after injection(24). Because only a small percentage of the spleen cells fromsuch mice have slg specific for SRBC, it seems likely thatlym-phokines induced by immunization with SRBC can cause B cellsto express a TRF receptor, just as interleukin 1 may induce Tcells to express an interleukin 2 receptor (25, 26).The authors thank Drs. Philip Lake and Ian Zitron for monoclonal

reagents, Drs. Irwin Scher, James J. Mond, and William E. Paul foradvice and review of the manuscript, Dr. Eva Suba and Mrs. JoanneSmith for technical assistance, and Ms. Clara Shields for editorial as-sistance and manuscript typing. This work was supported by the NavalMedical Research and Development Command Research Task no.MR041.20.01.0443 and Uniformed Services University of the HealthSciences Protocol no. RO 8308. The opinions and assertions containedherein are the private ones of the writers -and, are not to be construedas official or reflecting the views ofthe Defense Department, the NavyDepartment, or the"Naval Service at large.1. Sell, S. (1967) J. Exp. Med. 125, 289-301.2. Parker, D.. C. (1975) Nature (London) 258, 361-363.3. Scribner, D, J., Weiner, H. L. & Moorhead, J. W. (1978)J. Im-

munoL 121, 377-382.4. Sieckmann, D. G. (1981) ImmunoL Rev. 52, 181-210.5. Pure, E. & Vitetta, E. (1980) J. ImmunotL.125, 1240-1242.6. Mond, J. J., Sehgal, E., Kung, J. & Finkelman, F. D. (1981)J.

Immunol 127, 881-888.7. Chesnut, R. W. & Grey, H. M. (1981) J. ImmunoL 126, 1075-

1079.8. Ryan, J. J., Mond, J. J., Finkelman, F. D. & Scher, I. (1981) in

B Lymphocytes in the Immune Response, eds. Klinman, N., Mo-sier, D., Scher, I. & Vitetta, E. (Elsevier/North-Holland, Am-sterdam), Vol. 2, pp. 185-192.

9. Parker, D. C., Fothergill, J. J. & Wadsworth, D. C. (1979)J. Im-munoL 123, 931-941.

10. -Pur6, E., Isakson, P. C., Takatsu, K., Hamaoka, T., Swain, S.L., Dutton, R.. W., Dennert, G., Uhr, J. W. & Vitetta, E. S.(1981) 1. ImmunoL 127, 1953-1958.

11. Finkelman, F. D., Scher, I., Mond, J. J., Kung, J. T. & Metcalf,E. S. (1982) J. ImmunoL 129, 629-637.

12. Finkelman, F. D., Scher, I., Mond, J. J., Kessler, S., Kung, J.T. & Metcalf, E. S. (1982) J. ImmunoL 1.29, 638-646.

13. Takatsu, K., Tominaga, A. & Hamaoka, T. (1980) J. ImmunoL124, 2414-2422.

14. Tominaga, A., Takatsu, K. & Hamaoka, T. (1980) J. Immunot124, 2423-2429.

15. Lake, P., Clark, E. A., Khorshidi, M. & Sunshine, G. H. (1979)Eur. J. ImmunoL 9, 875-886.

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16. Mosier, D. E. (1969) J. Exp. Med. 129, 351-362.17. Cunningham, A. J. & Szenberg, A. (1968) Immunology 14, 599-

600.18. Finkelman, F. D., Kessler, S. W., Mushinskd, J. F. & Potter, M.

(1981) J. Immunol 126, 680-687.19. Zitron, I. M. & Clevinger, B. L. (1980)1. Exp. Med. 152, 1135-

1146.20. Schimpl, A. & Wecker, E. (1975) Transplant. Rev. 23, 176-188.21. Hilfiker, M. L., Moore, R. N. & Farrar, J. J. (1981)J. Immunol

127, 1983-1987.

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22. Vitetta, E. & Krolick, K. (1980)J. Immunot 124, 2988-2990.23. Delovitch, T. L., Watson, J., Battistella, R., Harris, J. F., Shaw,

J. & Paetkau, V. (1981) J. Exp. Med. 153, 107-128.24. Yaffe, L. J. & Finkelman, F. D. (1982) Ann. N.Y. Acad. Sci., in

press.25. Gillis, S. & Watson, J. (1981) ImmunoL Rev. 54, 81-109.26. Oppenheim, J. J. & Gery, I. (1982) Immunol Today 3, 113-117.

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