[CANCER RESEARCH 40. 2252-2256,
0008-5472/80/0040-OOOOS02.00
July 1980]
Indirect Rosette Microassay to Characterize Human Melanoma-associated
Antigens Recognized by Operationally Specific Xenoantisera1
Kohzoh Imai2 and Soldano Ferrone3
Department of Molecular Immunology, Scripps Clinic and Research Foundation, La Jolla, California 92037
ABSTRACT
MATERIALS
The indirect rosette assay which detects the interaction of
antibodies with target cells by their ability to rosette with sheep
red blood cells chemically coated with purified anti-immuno-
Cultured Cells and Murine Fetal Cells. Human melanoma
(BW5, M10, M14, M21, Colo 38, M51 ), carcinoma (D98, HT29,
MANO), sarcoma (S1), fetal fibroblast(IMR90),
lymphoblastoid
cell lines (Molt 4, 1301, Raji, RPMI 1788, RPMI 4098, RPMI
6410, RPMI 8866, Victor, WI-L2), and murine melanoma cell
line S91 were grown in Roswell Park Memorial Institute Medium
1640 with 10% fetal calf serum. These cell lines have been
described elsewhere (12). The identities of human cell lines
were periodically confirmed by HLA typing.
Polyclonal Xenoantisera to MAA and Cell Surface Markers
and Monoclonal Antibodies to HLA Antigens. Four types of
xenoantisera to MAA were used in this study. One (No. 6522)
was from a rabbit immunized with cultured melanoma cells
mixed with a xenoantiserum to cultured human lymphoid cells.
After extensive absorption with cultured human lymphoid cells,
the xenoantiserum reacted with melanoma cells and with other
types of tumor of nonlymphoid origin (12). Characterization of
the xenoantiserum by indirect immunoprecipitation
showed
that it contained one antibody population reacting with an
antigenic structure (M.W. 240,000) which has been detected
thus far only on melanoma cells and also one antibody popu
lation reacting with an antigenic structure (M.W. 94,000) ex
pressed both on melanoma cells and on other human tumors
of nonlymphoid origin (3). Xenoantiserum 8986 was from a
rabbit immunized with hybrids derived from the fusion of human
melanoma cells with murine fibroblasts; following absorption
with cultured lymphoid cells, the xenoantiserum contained a
major antibody population which immunochemical
studies
showed to react with the above-mentioned M.W. 94,000 com
ponent (3). Xenoantiserum 9456 came from a rabbit immunized
with MAA solubilized from cultured melanoma cells with 3 M
KCI and partially purified by affinity chromatography on lentil
lectin immunoadsorbents. The major antibody population in
this xenoantiserum reacted with the above mentioned M.W.
240,000 antigenic structure (3). Xenoantiserum 9446 came
from a rabbit immunized with MAA isolated from spent medium
of cultured melanoma cells by affinity chromatography on ricin
immunoadsorbents.
The major antibody population in this
xenoantiserum reacted with a M.W. 94,000 antigenic structure
(3). The xenoantisera to MAA used in this study had been
absorbed previously with pooled cultured B-lymphoid cell lines
to remove antibodies to HLA antigens and to human speciesspecific antigens.
A specific xenoantiserum to human ß2-fiwas produced in a
cow immunized with purified urinary ß2-fi(16). Rabbit antisera
to common determinants of HLA-A.B and HLA-DR antigens
have been prepared and characterized as described (20, 22).
The monoclonal antibodies 28 to framework determinants of
HLA-A.B antigens and 513 to framework determinants of HLADR antigens were prepared using the hybridoma technique (9)
globulin antibodies or with protein A from Staphylococcus
aureus has been used to analyze xenoantisera to human mel
anoma-associated antigens. The test has been developed as a
microassay and performed in microtiter plates, thus facilitating
the screening of large numbers of samples. When modified as
an inhibition assay, the assay has been successfully used (a)
to compare the specificities of xenoantisera elicited with cul
tured melanoma cells, hybrids derived from the fusion of cul
tured human melanoma cells, and murine fibroblasts and mel
anoma-associated
antigens purified by biochemical proce
dures and (b) to investigate the relationship of melanomaassociated antigens with /?2-microglobulin and HLA antigens
on the membrane of melanoma cells.
INTRODUCTION
Human melanoma cells express various types of membranebound antigens (5, 6), which have been characterized with
autologous (17), allogeneic (4, 10, 15), and xenogeneic (12,
18) antisera using a variety of assays (5). In our studies of
MAA4, we have elected to use serological assays since they
are simple and rapid and they allow the screening of a large
number of samples in a limited time. Binding assays have been
preferred to lytic assays to avoid the interference of changes
in susceptibility to lysis of target cells on the outcome of the
reaction. The aim of this study is to describe an indirect rosette
microassay to characterize xenoantisera to MAA and to inves
tigate their spatial relationship with other surface markers on
cell membrane. The test, which detects binding of antibodies
to melanoma cells by their ability to rosette with sheep erythrocytes chemically coated with anti-immunoglobulin antibodies
or with protein A from Staphylococcus aureus, is performed in
microtiter plates, thus facilitating the handling of large numbers
of samples.
' This work was supported by USPHS grants. This is Publication
1989 from
Scripps Clinic and Research Foundation.
2 Visiting investigator from Sapporo Medical College (Japan) and recipient of
a USPHS International Research Fellowship (1 F05 TW02817-01). To whom
requests for reprints should be addressed.
1Recipient of an American Heart Association Established Investigatorship
Award.
AThe abbreviations used are: MAA. melanoma-associated antigens; /32-/i. ßimicroglobulin; SPA. protein A from Staphylococcus aureus; SPA-E, sheep erythrocytes coated with protein A from S. aureus; Ab-E, sheep erythrocytes coated
with anti-immunoglobulin antibody; MEM, minimum essential medium.
Received December 3. 1979; accepted April 10. 1980.
2252
AND METHODS
CANCER
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Rosette Microassay and MAA
and characterized as described elsewhere (13, 21).
Coating of Sheep Erythrocytes with Protein A from SPA
and with Purified Antibodies to Immunoglobulin. SPA was
purchased from Pharmacia Fine Chemicals, Piscataway, N. J.
Antibodies to rabbit immunoglobulin and to mouse immunoglobulin were purified from goat anti-rabbit immunoglobulin
antisera and rabbit anti-mouse immunoglobulin antisera by
affinity chromatography on immunoadsorbent columns. SPA
and purified antibodies to immunoglobulin were coupled to
sheep erythrocytes by using the chromium chloride method
(14). Briefly, 1 volume of packed washed erythrocytes was
incubated with 1 volume of chromium chloride solution [0.1%
(w/v) in 0.15 M NaCI] and 1 volume of SPA (1 mg/ml) solution
or purified antibodies (1 mg/ml) for 5 min at 23°.SPA-E or AbE then were washed 3 times with 0.15 M NaCI and stored for
up to 7 days at 4°.
Indirect Rosette Microassay. Twenty /il of target cell sus
pensions (5 x 106/ml) were incubated with 20 fi\ of serial
80
320
1280
Dilution
Chart
1. Comparison
80
320
1280
Reciproca
of the efficiency
of SPA-E
and
Ab-E
in detecting
the
binding of rabbit MAA antibodies
to cultured melanoma
cells. Cultured melanoma
cells M21 and BW5 were incubated
with anti-MAA
Xenoantiserum
8986 (O. •)
or with normal rabbit serum (D, •)and then rosetted
with SPA-E (O. D) and AbE (••)
10CK
dilutions of antisera in microtiter plates (Microtiter U plates;
Cooke Laboratory Products, Alexandria, Va.) at 4°for 60 min.
80-
Then, the sensitized cells were washed 6 times with MEM and
pelleted by centrifugation at 200 x g at 4°for 5 min. Twentyfive /il of a 2% suspension of Ab-E or SPA-E then were added,
and the plates were centrifuged at 200 x g for 4 min at 4°.
The pellets were resuspended by vigorous pipetting and
stained with 25 /il of toluidine blue solution (0.1% in 0.15 M
NaCI). One drop of the suspension on a slide was examined
under a light microscope at 800 x g for resetted and unrosetted cells. A cell was considered rosetted when firmly sur
rounded by at least 5 Ab-E or SPA-E. Two hundred consecutive
cells were scored, and the percentage of rosettes was deter
mined.
In the rosette inhibition assay, 0.2 x 106 target cells were
incubated with 20 /il of an antiserum for 60 min at room
temperature. Where indicated, the cells were washed 3 times
with MEM and incubated with 20/il of a "sandwich" antiserum
(goat anti-rabbit immunoglobulin antiserum) for 30 min at 37°
to induce capping of antigens. After 3 washings with MEM, the
cells were incubated with an anti-MAA antiserum, and the
rosette assay was continued as described above.
Absorption. Xenoantisera were incubated with absorbing
cells at room temperature for 60 min.
RESULTS
Indirect Rosette Microassay. Preliminary studies compared
the efficiency of SPA-E for detecting the binding of xenoantibodies to melanoma cells with that of Ab-E. No significant
difference was found between the titration curves of an antiMAA xenoantiserum performed with the 2 types of indicator
cells, although at each dilution of the antiserum the percentage
of rosettes obtained with SPA-E is slightly greater than with AbE (Chart 1). With both types of indicator cells, the reproducibility of the indirect rosette microassay is high, as indicated by
the fact that titration curves of anti-MAA xenoantisera per
formed on different days are almost superimposable (Chart 2).
However, there is a certain degree of variation in the percent
age of rosettes detected with Ab-E when the value is lower
than 50%. The rosettes formed between antibody-sensitized
melanoma cells and SPA-E or Ab-E are stable up to 4 days of
JULY
»8
320
1280
80
320
1280
Dilution (Reciprocal)
Chart 2. Reproducibility of the indirect rosette microassay performed with
SPA-E and Ab-E. Titration curves of Xenoantiserum 8986 with cultured melanoma
M21 cells were performed on 5 days, using SPA-E (left) and Ab-E (right) as
indicator cells. •.
reactivity of melanoma cells with normal rabbit serum
storage at 4°,which eliminates the need to read the reactions
at the end of the centrifugation.
Reactivity of Xenoantisera to MAA with a Panel of Tumor
Cell Lines in the Indirect Rosette Microassay. The xenoanti
sera to MAA were tested with a panel of tumor cell lines which
included human melanoma cell lines, human B- and T-lymphoid
cell lines, human carcinoma cell lines, human sarcoma cell
lines, human fetal fibroblasts, and the murine melanoma cell
line S91. All the anti-MAA xenoantisera reacted with all of the
human melanoma cell lines tested with a titer of at least 1:80;
none of the xenoantisera reacted with the human lymphoid cell
lines and with the murine melanoma cell line S91. Furthermore,
Xenoantisera 6522, 8986, and 9446 reacted with carcinoma
and sarcoma cell lines, while Xenoantiserum 9456 did not
react with these types of tumor cell lines. Representative results
of the patterns of reactions obtained are shown in Table 1. Two
lines of evidence indicate that the reaction detected with the
indirect rosette microassay is specific. Target cells incubated
with normal rabbit serum did not rosette with Ab-E and SPA-E
(Chart 1). Furthermore, melanoma cells did not rosette with
SPA-E or Ab-E following incubation with anti-MAA xenoantisera
from which antibodies had been absorbed with melanoma cells.
Representative results are shown in Chart 3.
Rosette Inhibition Assay to Compare the Specificity of
1980
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2253
K. Imai and S. Ferrane
Anti-MAA Xenoantisera and to Investigate the Relationship
of MAA with Other Surface Markers. The rosette inhibition
assay was used to compare the specificity of anti-MAA xenoantisera produced with cultured melanoma cells, hybrids be
tween human melanoma cells and murine fibroblasts, and
purified MAA. In this test, cultured melanoma cells were first
sensitized with a given anti-MAA xenoantiserum; following re
distribution of antigens by incubation with a goat anti-rabbit
immunoglobulin antiserum, the cells were tested with another
anti-MAA xenoantiserum in the indirect rosette microassay.
SPA-E were used as indicators since SPA does not bind to
goat immunoglobulin (8). Representative results are shown in
Table 2. Melanoma M21 cells treated with Xenoantisera 6522
and 9446, both of which contain antibodies to a M.W. 94,000
structure expressed on tumor cells of nonlymphoid origin,
showed a significantly reduced reactivity with Xenoantiserum
8986 which contains antibodies to the M.W. 94,000 compo
nent; treatment of melanoma M21 cells with Xenoantiserum
9456, which contains antibodies to a M.W. 240,000 compo
nent detected only on melanoma cells did not affect the reac
tivity of the melanoma cells with Xenoantiserum 8986. Con
versely, melanoma M21 cells treated with Xenoantiserum 8986
under capping conditions lost their reactivity with Xenoanti
serum 9446 but did not have altered reactivity with Xenoanti
sera 6522 and 9456. These results indicate that Xenoantiserum
8986 reacts with the same antigenic structures recognized by
Xenoantiserum 9446 and by one of the 2 antibody populations
present in Xenoantiserum 6522, but not with those recognized
by Xenoantiserum 9456.
The rosette inhibition assay can also be performed by taking
advantage of the specificity of the anti-immunoglobulin
anti
bodies bound to sheep RBC, if the antisera being used have
been elicited in different animal species. This approach was
used to investigate the spatial relationship of MAA with /?2-ju
and HLA-A.B and HLA-DR antigens, using rabbit anti-MAA
antisera, cow anti-human ß2-nantisera, and murine monoclonal
antibodies to framework determinants of HLA-A.B and HLA-DR
100
60
BWS
:
MIO
\
£ 20
320
DILUTION
320
20
RICIPROCAI
Chart 3. Reactivity of Xenoantiserum 8986 with the cultured melanoma cells
BW5. M10, M14, and M21 after absorption with melanoma cells Colo 38 (•),
M10 (•)and M21 (A). O, reactivity of the unabsorbed xenoantiserum with the
melanoma cell lines.
Table 2
Comparison of the specificity of anti-MAA Xenoantiserum
Xenoantisera 6522, 9446. and 9456
withcSensitization
Resetted
8986 with that of
of ro
settes"1009278999682936
with3MEMMEMMEMMEM652294469456898689868986Capped
serum8986652294469456898689868986652294469456Tite
withGtaRblg"GtaRblgGtaRblgGtaRblgGtaRblgGtaRblgGtaRblgGtaRblgGtaRblgG
Table 1
Reactivity of an xenoantiserum to MAA solubilized from human cultured
melanoma cells
Xenoantiserum 9456 was from a rabbit immunized with MAA solubilized from
cultured melanoma cells by 3 M KCI and purified by affinity chromatography on
lentil lectin.
ofTumor
typeMelanomaCultured
% of rosettes at titer
lineM21M51RajiRPMI
1:16072
4352
30<1
B-lymphoidcellsCultured
788RPMI
1
4098RPMI
8866VictorWI-L2Molt
<1<1
<1<1
<14
<18
<1<1
<1<1
T-lymphoidcellsCarcinomaSarcomaFetal
41301D98HT29MANO81IMR901:20928651220293217753292813201:409071<!48182010<!<119<14<1141:80
<1<1
<14
1M21 cells (0.2 x 106) were incubated with 20 jil of MEM or xenoantiserum
at 4°for 60 min.
6 M21 cells sensitized with anti-MAA antibodies were incubated twice with
20 ftl of goat anti-rabbit immunoglobulin antiserum at 37°for 15 min.
c M21 cells after being capped were washed 3 times with MEM and then used
<1<1
<1<1
<1<1
<1<1
fibroblastCell
2254
<1
as targets in the indirect rosette assay.
" Mean value of duplicate experiments.
B GtaRblg. goat anti-rabbit immunoglobulin.
CANCER
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VOL.
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Rosette Microassay and MAA
antigens. Melanoma M21 cells coated with cow anti-human ß2fi antisera did not differ from melanoma cells incubated with
normal cow serum or with medium in their reactivity with rabbit
MAA Antiserum 8986 in the indirect rosette assay using sheep
erythrocytes coated with goat anti-rabbit immunoglobulin anti
bodies as indicators^results not shown). Furthermore, coating
of melanoma M51 cells with the rabbit anti-MAA Antiserum
6522 did not affect their reactivity with the anti-HLA-A,B (An
tibody 28) and anti-HLA-DR (Antibody 513) monoclonal anti
bodies in the rosetting assay using sheep erythrocytes coated
with rabbit anti-mouse immunoglobulin antibodies (Table 3).
These results indicate lack of close spatial association between
MAA and either ß2-fi,HLA-A.B, or HLA-DR antigens.
goats, ensuring a low background reaction level even if the
capping of the antigens has been incomplete.
The results of the rosette inhibition assay indicate that MAA
recognized by xenoantisera are not associated with /82-fi and
HLA antigens. This finding parallels similar observations by
Carey ef al. (1) using alloantisera and by ourselves using
immunochemical approaches.5 These data together argue
against the possibility that serologically detectable MAA are
structurally similar to HLA antigens or are modified HLA anti
gens (11, 19) and is in agreement with our previous finding
that MAA are not coded for by chromosome 6 which carries
the major histocompatibility complex region (2).
ACKNOWLEDGMENTS
DISCUSSION
The results reported in this study indicate that the indirect
rosette microassay which has already been successfully used
to detect HLA allo- and xenoantibodies (7) is a useful technique
for the serological characterization of anti-MAA xenoantisera.
When the test is performed in microtiter plates, only small
amounts of reagents are required and large panels of sera can
be screened easily. Sets of 96 samples can be handled as a
single unit during the washing procedures, and resetted cells
stored at 4°can be read up to 4 days after formation. Further
The authors wish to acknowledge the excellent technical assistance of June
Melin and Giorgio Brigato, the secretarial assistance of Dean Trinko, and the
editorial assistance of Andrea Rothman.
REFERENCES
1. Carey. T E., Lloyd, K. O.. Takahashi. T., Travassos. L. R., and Old. L. J. AU
cell-surface antigen of human malignant melanoma: solubilization and partial
characterization. Proc. Nati. Acad. Sei. U. S. A.. 76. 2898-2902.
1979.
2 Curry, R. A., Quaranta. V , Pellegrino, M. A., and Ferrane. S Serologically
detectable human melanoma associated antigens are not genetically linked
to HLA-A and B antigens. J. Immunol.. Õ22 2630-2632.
1979.
3. Ferrane, S., Imai, K., McCabe. R. P., Molinaro, G. A., Galloway, D. R., and
Reisfeld, R. A. Production, characterization
and use of xenoantisera to
more, the rosette microassay, equal in sensitivity to the radioimhuman melanoma associated antigens. In: S. Rosenberg (ed.), Serologie
Analysis of Solid Tumor Antigens. New York: Academic Press Inc.. in press,
munometric antibody-binding assay (3), is reproducible and
1979.
efficient as an inhibition test for comparing the specificity of
4. Ferrane, S., and Pellegrino, M. A. Cytotoxic antibodies to cultured melanoma
cells in the sera of melanoma patients. J. Nati. Cancer Inst., 58 1201different xenoantisera and in determining the spatial relation
1204, 1977.
ship of cell surface structures.
5. Ferrane, S., and Pellegrino, M. A. Serological detection of human melanomaThe 2 types of indicators used, SPA-E and Ab-E, did not
associated antigens. In: R. B. Herberman and R. Mclntire (eds.), Immunodiagnosis of Cancer, pp. 558-632. New York: Marcel Dekker. Inc.. 1979.
significantly differ in sensitivity to detect interactions of anti
6. Hellstram. K. E., and Brawn, J. P. Tumor antigens. In: M. Sela (ed.). The
bodies with target cells; however, SPA has the advantages of
Antigens. Vol. 5, pp. 1-82 New York: Academic Press, Inc., 1979
being commercially available as well not being reactive with
7. Indiveri. F., Wilson. B. S.. Pellegrino. M. A., and Ferrone, S. Detection of
human histocompatibility (HLA) antigens with an indirect rosette microassay.
immunoglobulin from certain species (e.g., goat) and with cer
J. Immunol. Methods, 29. 101-109. 1979.
tain immunoglobulin classes and subclasses. Therefore, in
8. Kessler. S. W. Cell membrane antigen isolation with the staphylococcal
rosette inhibition assays, SPA-E are useful indicator cells when
protein A-antibody adsorbent J. Immunol.. 117: 1482-1490.
1976.
the "sandwich"
anti-immunoglobulin
antiserum is raised in
9. Kohler. G., and Milstein. C. Continuous cultures of fused cells secreting
antibody of predefined specificity Nature (Lond.), 256 495-497. 1975
10. Lewis, M. G., and Phillips, T. M. The specificity of surface membrane
immunofluorescence
in human malignant melanoma Int. J Cancer. 10
Table 3
105-111, 1972.
Lack of spatial relationship between HLA antigens and serologically detectable
11. Malley, A , Burger, D. R.. Vandenbark, A. A.. Frikke. M.. Finke. P., Begley.
MAA. as determined by the rosette inhibition assay
D., Acott. K.. Black. J.. and Vetto. R. M. Association of melanoma tumor
antigen activity with ft-microglobulin.
Cancer Res., 39. 619-623, 1979.
Resetted with
12. McCabe. R. P.. Quaranta. V.. Frugis, L., Ferrone, S., and Reisfeld, R. A. A
%of
radioimmunometric antibody binding assay for the evaluation of xenoantisera
Titer
rosettes
Antiserum
Sensitizaron with
to melanoma associated antigens. J. Nati. Cancer Inst.. 62 455-463. 1979.
Normal rabbit
serum(1:5)8492°
13. Molinaro, G. A., Quaranta, V., and Ferrone. S. Characterization of the
specificity of DR monoclonal antibodies. Transplant. Proc., 11:1748-1750.
(1.5)6522d(1:5)Normal
1979.
14. Molinaro. G. A.. Sato. K., Paque. R. E., and Dray, S. Rosetting of antibodysensitized tumor cells with anti-immunoglobulin
antibody-coated erythro
cytes by a new method for detecting antigens on cells. Cancer Res., 35.
2213-2217.
1975.
serum(1:5)861
rabbit
15. Morton, D. L., Malmgren, R A.. Holmes, E. C.. and Ketcham. A S. Demon
(1:5)6522(1:5)2828282828285135135135135135131:11:101:11:101:11:101:11:101:11:101:11:107763268785878592097552
2e
stration of antibodies against human malignant melanoma by immunofluo
rescence. Surgery. 64: 233-240, 1968.
16. Pellegrino, M. A.. Belvedere. M., Pellegrino, A. G., and Ferrone, S. B
peripheral lymphocytes express more HLA antigens than peripheral lympho
cytes. Transplantation (Baltimore). 25. 93-95, 1978.
M51 cells (0.2 x 106) were incubated with 20 fil of each reagent at 4°for
17. Shiku, H., Takahashi, T., Oettgen. H. F.. and Old, L. J. Cell surface antigens
60 min; following 3 washings, the cells were tested with Monoclonal Antibody 28
of human malignant melanojna. II. Serological typing with immune adherence
(anti-HLA-A, B) or Antibody 513 (anti-HLA-DR) in the indirect rosette microassay
assays and definition of twoi>ew surface antigens. J. Exp Med..144: 873using sheep erythrocytes coated with rabbit anti-mouse immunoglobulin antibod
881. 1976.
ies as indicators.
6 Mean value of duplicate experiments.
c Rabbit anti-HLA-A, B. C antiserum.
5 R. P. McCabe, F. Indiveri. S. Ferrone, and R. A. Reisfeld. Lack of association
d Rabbit anti-MAA antiserum.
of serologically detectable human melanoma associated antigens with /¡2-microe Rabbit anti-HLA-DR antiserum.
globulin. submitted for publication.
JULY
1980
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1980 American Association for Cancer Research.
2255
K. Ima/ and S. Ferrone
18. Stuhlmiller, G. M., and Seigier, H. F. Characterization of a chimpanzee antihuman melanoma antiserum. Cancer Res., 35. 2132-2137,
1975.
19. Thomson, D. M. P., Tataryn, D. N., O'Connor, R., Ranch, J., Friedlander, P..
J. Immunol., 122: 1967-1971,
1979.
21. Wilson, B. S.. Ng, A. K.. Quaranta, V., and Ferrone, S. HLA polyclonal and
monoclonal xenoantibodies: production, characterization and application to
Gold, P., and Shuster, J. Evidence for expression of human tumor-specific
antigens associated with /i2-microglobulin in human cancer and in some
colon adenomas and benign breast lesions. Cancer Res., 39. 604-611,
1979.
20. Wilson, B. S., Indiveri. F., Pellegrino, M. A., and Ferrone, S. Production and
characterization of DR xenoantisera: use for detection of serum DR antigens.
the study of HLA antigens. In: R. A. Reisfeld and S. Ferrone (eds.). Current
Trends in Histocompatibility. New York: Plenum Press, in press, 1980.
22. Wilson, B. S., Pellegrino, M. A., Reisfeld, R. A., and Ferrone, S. A simple
method for production of specific xenoantisera to human histocompatibility
(HLA-A,B,C) antigens. Transplant. Proc., 10: 741-743. 1978.
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Indirect Rosette Microassay to Characterize Human
Melanoma-associated Antigens Recognized by Operationally
Specific Xenoantisera
Kohzoh Imai and Soldano Ferrone
Cancer Res 1980;40:2252-2256.
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