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Two-dimensional isoelectrie focusing/SDS PAGE of the lentil binding and lentil nonbinding forms of gp 100 over a narrow pH range gave identical homogeneous spots with a pI of 5.1-5.5. Neuraminidase treatment of both forms of gp 100 resulted in a shift of approximately 0.6 of a pH unit towards the basic end of the gel. 11. Is Carbohydrate Involved in the Antigenie Site of gp 100? Treatment of cells with tunicamycin (20 !!g/ml) and examination of immunoprecipitates from extracts on SDS-PAGE resulted in the disappearance of gp 100 with the concomitant appearance of a molecule at 75K. This was precipitable using rabbit anti sera and a monodon al anti-ALL (Ritz et al. 1980). Digestion of cell extracts with a mixture of glycosidases also gave an immunoprecipitable band of lower molecular weight. This evidence plus the heat lability of gp 100 suggests that carbohydrate, while possibly constituting up to 25 % of the molecular weight, is not involved in the antigenic site. IV. The gp 100 on Normal Bone Marrow Cells Small numbers of cells in normal bone marrow react with anti-ALL (Greaves et al. 1978, 1980; J anossy et al. 1978; see also contribution by Greaves et al. in this book). Uninvolved bone marrow from three cases of children with rhabdomyosarcoma and two sampies of marrow from patients in remission from leukemia were metabolically labelIed with 35S-methionine and immunoprecipitates of cell extracts examined by SDS-PAGE. A glycoprotein with a molecular weight of 100,000 daltons identical to that seen in leukemias and leukemic cells lines was detected. Rabbit anti sera and monodonal (J-5) antibody precipitated a similar band (Fig. 2). Peptide mapping will be required to establish 111. Is gp 100 an Integral Membrane Protein ? As gp 100 is shed into the culture medium(s) and is also present in the sera of some leukemic patients (Kabisch et al. 1979; unpublished observations), it was of interest to establish whether or not it is an integral membrane protein. Incubation of Nalm-1 cells with the lipophilic photoactivatable reagent, hexanoyl diiodo N -( 4-azido-2-ni trophenyl)-tyramine (Owen et al. , to be published), was able to label Ia antigens but no labeling of gp 100 was evident. Charge shift electrophoresis (Helenius and Simons 1977) showed that the molecule possessed limited hydrophobicity, although this was no greater than that associated with the soluble proteins, bovine serum albumin, and lysozyme. Thus, although gp 100 possesses some hydrophobie regions, it is most likely not a true integral membrane protein. Subcellular fractionation of 125I1lactoperoxidase-Iabeled cells, however, showed that gp 100 is strongly associated with the plasma membrane and not released into the medium as is the case with known peripheral proteins such as fibronectin (Owen et al., to be published). b c d Fig. 2. SDS-PAGE of immunoprecipitates from 35S-methionine labeled bone marrow cells. a, rhabdomyosarcoma and rabbit anti-ALL; b, rhabdomyosarcoma and monoclonal anti-ALL (J-5); c, remission marrow and rabbit anti-ALL; and d, remission marrow and normal rabbit serum 327
complete identity of gp 100 from normal marrow cells with that of leukemic cells. This data and other associated phenotypic features (see Greaves et al. pp 296-304) indicate that expression of gp 100 is likely to be anormal gene product reflecting the "target" cell for malignant transformation and/or the level of maturation arrest commonly seen in ALL. References Graham JM, Hynes RO, Davidson EA, Bainton DF (1975) The loeation of proteins labelIed by the 125I-Iaetoperoxidase system in the NIL8 hamster fibroblast. Cell 4:353-365 - Greaves MF, Janossy G (1978) Patterns of gene expression and the eellular origins of human leukaemia. Bioehim Biophys Aeta 516: 193-230 - Greaves M, Janossy G, Franeis G, Minowada J (1978) Membrane phenotypes of human leukaemic eells and leukaemic eell lines: Clinical eorrelates and biological implieations. In: Clarkson B, Marks PA, Till JE (eds) Differentiation of normal and neoplastic hematopoietie eells. Cold Spring Harbor, New York pp 823-841-Grea- ves M, Delia D, Janossy G, Rapson N, Chessells J, Woods M, Prentiee G (1980) Aeute lymphoblastic leukaemia assoeiated antigen. IV. Expression on non-Ieukaemic 'lymphoid' eells. Leuk Res 4: 15-32 - Helenius A, Simons K (1977) Charge shift eleetrophoresis: Simple method for distinguishing between amphiphilie and hydrophilie proteins in detergent solution. Proe Natl Aead Sei USA 74:529-532 - Janossy G, Franeis GE, Capellaro D, Goldstone AH, Greaves MF (1978) Cell sorter analysis of leukaemia-assoeiated antigens on human myeloid preeursors. Nature 276: 176-178 -Kabiseh H, Arndt R, Beeker W-M, Thiele H-G, Landbeek G (1979) Serological deteetion and partial characterization of the eommon-ALL-eell associated antigen in the serum of eALL-patients. Leuk Res 3: 83-91 - Owen MJ, Knott JCA, Crumpton MJ (to be published) Labelling of lymphoeyte surfaee antigens by the lipophilie, photoaetivatable reagent, hexanoyl diiodo-N-( 4-azido-2-nitrophenyl)-tyramine. Bioehemistry - Ritz J, Pesando JM, MeConarty JN, Lazarus H, Schlossman SF (1980) A monoclonal antibody to human aeute lymphoblastic leukaemia antigen. Nature 283:583-585 - Sutherland DR, Smart J, Niaudet P, Greaves MF (1978) Acute lymphoblastie leukaemia assoeiated antigen. H. Isolation and partial eharaeterisation. Leuk Res 2:115-126 328
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Haematology and Blood Transfusion 2
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Dr. Rolf Neth, Universitäts-Kinder
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Participants of the Meeting Aaronso
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Kurth, R, Friedrich-Miescher-Labora
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Wilsede Scholarship Holders (Grante
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Preface Gut ist eine Lehrart, wo ma
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Personal and scientific discussion
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Acknowledgement We should like to t
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Wilsede, June 21 1978 Memorial Trib
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limiting benign lymphoproliferative
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this most unlikely, however (for re
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longer subject to the same control
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Bregula U, Wiener F, Harris H (1971
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fresh lymph node biopsies from ten
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Fig. 3. Binucleate mitosis in an ob
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y Fig. 4. Schematic diagram of post
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than that among patients with Stage
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Tests of binding affinity, agglutin
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N Engl J Med 297: 963-968 - Green I
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Clinical Aspects
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"ring chromosome". "Mar" is marker,
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191 chromosomally abnormal patients
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tumors of both African and non-Afri
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Haematology and Blood Transfusion V
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Retrospective group 26/93 evolved l
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Treatment
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advantage in terms of duration of f
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leukaemia. An analysis of 168 patie
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11. Treatment Program Chemotherapy
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efractory congestive heart failure
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period (Sequence II) was gene rally
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children with a multiple drug proto
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DAYS 1 5 10 15 20 25 30 35 , I I I
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Biologically Drug Outcome fit" sens
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may counterbalance the potential be
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Ht (0--0) 50 40 30 20 10 0 Platelet
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10 7 ,-----------------------------
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11. Haematological and Biochemical
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0\ 0\ INTERFERON ... 10 3 rl E "'
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("pre-B" phenotype) has been descri
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human bone marrow cells exhibit the
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10 9.,0 MBKCC .. .,4 B prot:oool 1'
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demann W, Büehner T, Arlin Z, Gee
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-...l 00 a-tLDRENS CAt«:ER S TU>Y
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prognostic characteristics under in
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significantly higher in HR} (14.5%,
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normal or elevated immunoglobulin l
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References Andreeff M, Darzynkiewic
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R GROUP 1 ( 1970 - 1971 ) 17 PTS. R
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Table 3. Influence of initial featu
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LIJ (f) a.. < -1 LIJ Q: u. 0 >- I-
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me nt indicated that additional the
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42:2123-2134 - Chessells JM, Cornbl
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Baum et al. 1979). This study was b
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1. 0 '--'1--" 0.9 e: o .- VI VI E C
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e: o e: 1. 0 ,.--...-- 1'"'\ ......
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In conclusion, we can state that th
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Haematology and Blood Transfnsion V
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INDUCTION VCR + PRED + ASNASE (wk 3
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SCHED.INT 1 MTX • ! Adria MTX •
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~ looh---------ooooc>----o---c>----
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Consoll- Irra .tlon t) .tlon Indu
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Highrisk Standard risk Table 1. Cli
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Cumulo/ive comp/el9 remission 1.0 .
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10° 8 = 6 ö 4 Non T Cell .~ :ö
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% SURVIVAL 100 ~ ~II 0 80 60 40 - -
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a) Percentage of donor cell mitoses
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Table 5. Results of bone marrow tra
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...... w N Table 7. Clinical result
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agent had to be added and that in t
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C. Chronic Myelogenous Leukemia The
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Patients with a suitable donor rece
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No. Recurrent Leukaemia Other death
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followed by dialysis against isoton
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HS, HL- Heme >_ 40 0 HS I ;:. =====
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knowing at present. These genes may
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esistance to antifolates. In Vitro
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a DNA probe specific for the gene t
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translocation between chromosomes 9
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Table 1. Subdivision of 1827 Myelo-
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NORMAL CELL ®
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some change of the primary type, in
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F. Transformation of Mouse Bone Mar
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were carried out with 3H-MTX in the
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the lysozyme cDNA prepared from ovi
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et al. 1976). The BJAB cellline in
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dalton bands corresponding to light
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1979). The EBV-carrying cell lines
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producer lines (DAUDI and P3HR-1),
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G, Giovanella B, Westman A, Stehlin
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Dc;bt CT + i J.I9 Raji; 1. i .. 100
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c. Discussion During infectious mon
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C. Nonepisomal Viral DNA We have us
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A 260 3.0 A B 1670 70 N2 2.0 c: E .
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vivo. Nature 260:302-306 - Kirk JTO
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Fig. 1. a Polyacrylamide gelelectro
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1979). The lymphocyte subpopulation
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lysin 0 antigens are found, and mix
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Pope JH (1978) Long-term T cell-med
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Haematology and Blood Transfnsion V
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Charaeteristie Morphologie maturati
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after TPA-induction (Table 3). The
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PA (1978) Reactivity of a rabbit an
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Blood Monocyte. B lood Monocytes 1-
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and nonhistone proteins can serve a
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The findings of the evaluation of s
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Fig. 3. Tumor grawth of L 428 cells
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Cell biological and Immunological A
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consideration. Sanel (1973) obtaine
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endotoxin serum, were used. A parti
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monocytic leukemia cell line in cul
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B. Isolation, Characterization and
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don al hemopathies generally displa
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nitors. Blood Cells 6:595-607 - Min
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\I) ....I ....I IU Jl 200 r 100 90
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their haemopoietic cells' ability t
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normal normal ALL ALL ALL persons p
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64.5 { V)
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References Biermann E, Neth R, Gros
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ehambers, the growth pattern observ
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References Anderssen LC, 10kinen M,
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REH CELL UNE TO CALLb~Ö ~q~ V.M. c
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the macrophage series in which the
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Table 1. Production of factor depen
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Page 289 and 290: Table 1. Characteristics of the adh
Page 291 and 292: Fig. 3. A human marrow culture at 6
Page 293 and 294: 300 250 • HYPERPROLIFERATIVE PATT
Page 295 and 296: Fig. 9. Nonadherent population from
Page 297 and 298: The cobblestone areas were often no
Page 299 and 300: L (1976) Induction of colony format
Page 301 and 302: B. Materials and Methods J. Tissue
Page 303 and 304: Fig. 2. Ultrastructural appearance
Page 309 and 310: Table 1. Monoclonal antibody reacti
Page 311 and 312: al. 1979; Janossy et al. 1979) is e
Page 313 and 314: to which this is an exact replica o
Page 315 and 316: inger JL (1976) Distribution of la-
Page 317 and 318: I region antigens were found to be
Page 319 and 320: have been reported to be present on
Page 321 and 322: V. FunctionalAssays Assays for PHA
Page 323 and 324: Functional studies of UCHT1 separat
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Page 329 and 330: with immunohistochemieal methods (L
Page 333 and 334: can be dissected by cloning. Utiliz
Page 335 and 336: Table 1. Origin and marker profile
Page 337: Haematology and Blood Transfusion V
Page 341 and 342: Table 1. Reaction of single anti se
Page 345 and 346: media. Following 24-h incubation, c
Page 347 and 348: nonleukemic eells (Greaves 1979). L
Page 351 and 352: Fig. 2. Cytocentrifuged smears of b
Page 353 and 354: Modification of amino-groups of hum
Page 355 and 356: disturb this balance in the directi
Page 357 and 358: The majority of human blood lymphoc
Page 359 and 360: of the EBV infected B cells. Howeve
Page 361 and 362: and immunoglobulins. J Immunol 120:
Page 363 and 364: " K 10 • j ~ ALL RNlL AL wtfh les
Page 365 and 366: tion akuter Leukämiezellen in "spo
Page 367 and 368: D. Results and Discussion J. Acute
Page 369 and 370: selectively responsive to different
Page 373 and 374: l1J ~ 100 ~ Q. ::) l1J Z 0 ~ z >- .
Page 375 and 376: Table 2. Growth inhibition of S49 e
Page 377 and 378: Reverse Infectious ASC/2X1Q6 transc
Page 381 and 382: munized animals to 1316 tumor cells
Page 383 and 384: Haematology and Blood Transfnsion V
Page 385 and 386: Fig. 3. Expression of retroviral gp
Page 387 and 388: pg70. J Exp Med 143: 151-166 - McGr
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Table 1. Expression of tumor antige
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Virological and Molecularbiological
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Table 1. Oncogenic properties of re
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also Fig. 1). It followed that the
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whether this sequence is related to
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zed in infected cells argue that th
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a specific viral transforming prote
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detect viral RNA as the only charac
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Eisen H (1980) Myeloproliferate vir
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p,60- -34K - 1 2 3 4 Fig. 1. In vit
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.. p - • a'T ... I .t'l Fig. 4. D
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Fig. 2. Photomicrographs of NY68 in
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DEAE (o(lJmn 'liI GI -2 2 o e s 1
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vity was deterrnined. For endogenou
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pr 180 - - - P 12/15 - A B c D E F
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85,- 66- -27 - -66 40- 27- -1'9 19-
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= 0.5 o E Q. o ILI 0.4 (J) « ILI .
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Table 1. Oncogenic potential of avi
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100 III GI c: o "8 80 > w
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Expt. No. Infecting virus Proportio
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Haematology and Blood 'fransfusion
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sequences, provided its gag portion
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77:3009-3013 - Hu SSF, Moscoviei C,
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RI 2.2 md and Hind III 2.6 md leuke
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RELATIONSHIP OF ENDOGENOUS AND EXOG
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E ........ E Cl. U E :::l .... Q) (
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References Astrin S (1978) Endogeno
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the same animal were examined (e.g.
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I) 'IQ a \0 - CL -- Fig. 2. Restrie
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~ ..................... ~ Cell 3' 5
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a :Sa,C! + . ~ 111 i' ( " '~ .J. b
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Table 1 Sampie Tissue TS fragment"
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a b Sac I / rep ._-_.-_ K J nl rep
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Haematology and Blood 'fiansfusion
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Osteopetrosis and osteosarcomas occ
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Lymphoma cellline Table 1. T and B
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Table 2. In vivo growth and oneogen
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Table 1. Transforming aetivity of c
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Table 3. Transformation aetivity of
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- Shoemaker C, Goff S, Gilboa E, Pa
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indieate a shifting of target eell
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Table 2. Expression of Jl and Sourc
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~ i ,..1---------,,':::::::::::::::
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fragment of pBR322 DNA, a 3.0-kbp S
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Mak TW (1979) Proc Natl Acad Sci US
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Table 1. Transformation of 3T3 cell
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inserts in the genome of rapidly tr
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pulations, it seemed likely that th
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contains antigens which react with
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le of reacting in sensitive radioim
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gic approach. In: Hiatt HH, Watson
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12 Eco R [ Fig. 1. Hybridization pa
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10 - '1~ )( -~ S:! E A U I o 1 2
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Haematology and Blood 'fransfusion
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cultures. These cells did not conta
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indication of malignant T-cells in
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Fig. 3. Thin-section electron micro
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Table 3. Lack of detectable related
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J. HTLV Was Present in the Primary
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specific signals into nonspecific s
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Table 2. Distribution of HTLV-relat
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u 10 ~ 20 ~ :c 30 a 40 o Fig. 1. Ki
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SSV TrS-gp labeled effectively with
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60 HFF/SSV cDNA CELL RNAs: 0 HF/SSV
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Antisera a Table 1. Immunofluoresce
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Haematology and Blood 'ftansfusion
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y competition RIA yielded virtually
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SiS'V' gp70: ANTIGEN, ANTI BODY, IM
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type-C virus p30 antigen in cells f
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c. Results The fractionated whole-b
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induced antibodies as well as exper
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- Fig. la-f. Retravirus particles p
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eplication of infecting murine leuk
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immunodeficiency, multic10nal lymph
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Bovine leukemia --, proteins of 499
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Human T- cell - -, characterization
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RNA -, of tumor virus 439 Rous sarc
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Haematology andBlood Transfusion Su
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