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equilibrium in the sap, i.e., when the concentration of dye in the sap be-came constant over a period of several hours, then the concentration ofdye in the sap was 1/4 of that in the external solution when phosphatebuffers were used, and l/s with borate buffers.These results agree with those found for the species of Valonia from

Florida, V. ventricosa. The laws governing the penetration of thesedyes into different species of Valonia are essentially alike.This is a preliminary paper. Details will be published subsequently.2

Grateful acknowledgment is hereby made to the Bache Fund of theNATIONAL ACADEMY OF SCIENCES for a grant enabling the writer to carryon these experiments in the South Seas.

I Kindly identified by Dr. W. A. Setchell.2 Brooks, M. M. Protoplasma. In process of publication.

STUDIES ON MOUSE LEUKEMIA. VI. THE PREDOMINATINGCELL TYPE IN LINE I*

By JAMES S. POTTER AND MAURICE N. RICHTER

DEPARTMENT OF GENETICS, CARNEGIE INSTITUTION OF WASHINGTON, COLD SPRINGHARBOR, AND THE DEPARTMENT OF PATHOLOGY, COLLEGE OF PHYSICIANS AND SURGEONS,

COLUMBIA UNIVERSITY, NEW YORK

Communicated February 26, 1932

Lymphatic leukemia occurring spontaneously and regularly in miceof Strain C58'0 has been transmitted by inoculation to young mice ofthe same strain. This has led to the establishment of a series of trans-mission lines (designated by letters of the alphabet) each with its individualcharacteristics." Line I has been carried experimentally through 167transfers in mice of Strain C58 and has shown considerable constancyboth in the gross and microscopic pictures and in the interval betweeninoculation and death. Susceptibility to Line I has been confined tocertain strains of mice. A genetic analysis of the susceptibility of StrainC58 to this line has been reported elsewhere."6

All efforts thus far to separate an active agent causing this leukemiafrom the cellular component of the inoculated material have been un-successful.9 But the question still remains, whether the cells of thecharacteristic accumulations in the inoculated animals are descendants ofthe living cells introduced by inoculation or of cells originating in the host.The following report is a first step in the cytological approach to thisproblem.

Materials and Methods.-Animals of Strain C58 were taken at 24-hour

298 PROC. N. A. S.

PLATE 1 (Photomicrographs).

Fig. 1. Cells in periphery of follicle in a spleen 96 hours after inoculation.(Polar view mitotic figure in upper right.)

Fig. 2. Portion of perivascular accumulation in liver 72 hours after inoculation.Fig. 3. Infiltration of abdominal wall 96 hours after inoculation.Fig. 4. Perivascular accumulation in kidney 120 hours after inoculation.Fig. 5. Dividing cells in a germinal center of a cervical node from an uninocu-

lated mouse of Storrs-Little strain, in which spontaneous leukemia is not knownto occur. Cell type indicated by arrows morphologically similar to the activecells of Line I.

Fig. 6. Mitosis in peripheral blood of mouse inoculated with Line T leukemia.Smear preparation, Flemming's fixation and Wright's stain.

GENETICS: POTTER ANDA CHTER

intervals after intraperitoneal inoculation with a normal saline suspensionof minced spleen from previous inoculations, i.e., the 121st, 122nd, 123rdand 133rd transfers of Line I. In the last of these transfers animals werealso killed every four hours during the first twenty-four hour period afterinoculation. Tissues were taken at the same time from uninoculatedanimals as controls to show the activity of the lymphatic tissues of animalsin the same environment, of the same age and younger. Tissues from

.... 'f ..s ,.. .. ..

A ::*; s-s-|_.. ., :._

(.

*e iaN as: ,. 's .... ,^ . :::. ..... !: i'_ . ! .

TEXTFIGURE 1

a. Nuclear organization of infiltrating cellfrom sinusoid of a liver. Drawing made fromFeulgen preparation with normal color intensi-ties inverted to emphasize plasmasome. b.Cell from perivascular accumulation in kidney.Flemming's triple stain. c. Small lympho-cyte. d. Prophase in infiltrating cell type.e. Late prophase chromosomes.

(Camera lucida outlines a-d approx. X 2300,e approx. X 1900.)

paraffin and cut in serial sections 5

embryos were also examined.Animals were killed by sev-

ering the spinal cord. Fromeach mouse the followingmaterial was preserved: lymphnodes, spleen, liver, kidneys,lungs, genitals, abdominal walland any other tissue thatsuggested infiltration in thegross picture at autopsy. Alltissues were in the fixing fluidwithin twelve minutes afterthe death of the animal.

Several methods of fixationwere tried in the preliminarystages of this work, but onlytwo were used for routinepurposes. Flemming's fluidfor twenty-four hours at3-6°C., and Winge's'3 com-bination of picro-Carnoy fol-lowed by B-15 gave the bestresults. For special studiesthe Feulgen reaction wasused, as well as fixationand staining according toSchridde.3'7

Tissues were embedded into 7 micra in thickness. All

tissues fixed by routine methods were stained either with Haidenhain'siron-alum haematoxylin or with Flemming's triple stain. Part ofthe embryo material studied was stained with haematoxylin andeosin. Light green was used as a counter-stain following the Feulgenreaction.

Blood and tissue smears were made for staining with Wright's stain,as well as with the technique used by Wiseman14 for determining the

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GENETICS: POTTER AND RICHTER

relative age of lymphocytes. Smears were also fixed with Flemming'sfluid followed by staining with Wright's. Observations were madeon cells stained supravitallywith Janus green and neu-tral red. Sato's"2 peroxidasemethod for blood smears was >"t

Observations.-The accumu-lations of cells in the lym-phatic tissues and elsewhereappear to consist predomi- f 4 inantly of a large type cellof the lymphatic series. This AM.is true of all stages up to thedeathof the animal. A series ...iof measurements in sectionedmaterial placed the size rangeof the cell type between 9 and .

11 micra when rounded up.This cell type has a mod-

erate amount of cytoplasmcontaining many large, roundmitochondria (evidenced by Xsupravital, staining withJanus green as well as bythe Schridde section methods) 4which, with one or severalvacuoles, are usually locatednear to the nuclear mem-brane, and have been ob-served in a number of cases TEXTFIGURE 2to clump in a position near a. Portion of mesenteric node from anto the nucleolus. There may inoculated mouse after preparation by thebe a few azure granules or Feulgen technique. Green counter-stain fil-none at all. Observations tered in photographing resulting in faint nu-on basophilia as described by cleolar outlines and emphasizing chromatin

arrangement. (Photomic.) b. Blood smearWman ic

that ter from inoculated mouse showing negativemajprity of the cells were (arrows) peroxidase reaction. Cells showing

young. positive reaction are polymorphonuclears.The nucleus is usually ir- (Photomic.)

regular in shape. The posi-tion and size of the nucleolus and the arrangement of the chromatin maybest be demonstrated by the Feulgen technique, in which the nucleolus

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takes a brilliant green from the light green counter-stain in contrast tothe purple of the chromatin. The nucleolus is a large, elongated plas-masome, irregular in shape and adherent through most of its length tothe nuclear membrane (Textfig. la). During the "resting" conditionthe chromatin is dispersed in an open network with a tendency toclump about the nucleolus (Textfigs. lb and 2a). The size and positionof the plasmasome and the arrangement of the chromatin complementbecome identifying features of the cell type.

Cell division is found in every section of the areas affected. In someof the earlier stages of infiltration about fifty per cent of the cells involvedhave been observed in some active stage of division.

During the early prophase the spireme is coarse and is in the form ofloops radiating from a point adjacent to the plasmasome. The strandsof the loops usually adhere to the nuclear membrane, making it possibleto count the strands in optical section (Textfig. ld) and giving a strikingappearance to this stage. At the time of fragmentation of the spiremeand formation of the individual chromosomes the plasmasome has dis-appeared entirely.

Because of the tendency of the chromosomes to split precociously atmetaphase, late prophase was found to be the best time at which to makecounts of the chromosomes (Textfig. le). A series of fifty counts wasmade in areas of accumulated cells in different tissues and at variousstages of development following inoculation. Camera lucida outlinesof the chromosomes were made to facilitate counting and to insure ac-curacy. The nujmber in the figures examined was invariably forty, the2n number reported for the mouse by Masui,4 Cox,' Minouchi8 andothers. No effort has been made so far to study the morphology of theindividual chromosomes.No variation from normal metaphase and anaphase was observed except

in areas with much cellular degeneration. The abnormalities were sorare, however, that they are interpreted as being of no consequence inthe continuity of a cell type, but rather the result of abnormal physio-logical conditions in the degenerative areas. Abnormal mitosis is atypicalof most stages of this line of leukemia, and is not comparable in incidencewith the chromosomal aberrations reported by Ludford2 in the cells of atransplantable tumor of the mouse, or with variations reported by Winge'3in tar carcinomata.Abnormal "inclusion bodies" have not been observed so far. The

peroxidase reaction of the cell type is negative (Textfig. 2b).Observations in the developing liver of mouse embryos and in lymphatic

tissues in unweaned and maturing mice of different strains revealed cellsof a type morphologically identical to the cell type predominating in theareas of infiltration in the inoculated mice. In the infiltrated areas inter-

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mediate stages appear between the cell type described here and the normalsmall lymphocyte of the mouse.Summary.-A cytological description is given of the cell type pre-

dominating in accumulations in organs and tissues and resulting fromexperimental inoculation of Line I leukemia. The chromosome numberis forty, the normal for somatic cells of the mouse. Mitotic aberrationsare not present in significant numbers. None of the cytological methodsused differentiated between the cell type described and a morphologicallysimilar type found in normal mouse tissues.

* This investigation was aided by a grant from the Carnegie Corporation.1 Cox, E. K., J. Morph. and Phys., [1] 43, 45 (1926).2 Ludford, R. J., Report on Investigations, Imp. Cancer Res. Fund, 145-153 (1930).3 Lee, B., Microtomist's Vade Mecum, 9th Ed., Blakiston (1928).4 Masui, K., J. Coll. Agr., Imp. Univ. Tokyo, 8, 207 (1923).5 MacDowell, E. C., and Richter, M. N., J. Cancer Res., [3] 14, 434-439 (1930).6Ibid.,Proc. Soc. Exp. Biol. and Med., 28, 1012-1013(1931).7McClung, C. E., Microscopical Technique, Hoeber (1929).8 Minouchi, O., Jap. J. Zool., 1, 269 (1928).9 Richter, M. N., and MacDowell, E. C. (Abstract), Arch. Pathol., 9, 1299 (1930).10 Ibid., Proc. Soc. Exp. Biol. and Med., 26, 362 (1929).11 Ibid., J. Exp. Med., [4] 51, 659-673 (1930).12 Sato, A., and Yoshimatsu, S., Am. J. Dis. Child., 29, 301 (March, 1925).13 Winge, O., Zeit. f. Zellforschung Mikr. Anat., [4] 10, 683-735 (1930).14 Wiseman, B. K., J. Exptl. Med., [2] 54, 271-294 (1931).

A CRITIQUE OF RECENT QUANTUM THEORIES. IH

BY R. J. SEEGERTHE GEORGE WASHINGTON UNIVERSITY

Communicated March 2, 1932

The sine qua non of a physical theory is its ability to describe experi-ential facts accurately and uniquely. But the very first uses of thenew quantum mechanics hinted of doubts as to the satisfying of thisdemand. The special methods employed by Pauli and by Dirac in theirsolutions of the hydrogen atom were hardly general. The arbitrarynumber of degrees of freedom used to solve the rotator lacked definiteness.Indeed, the later discovery of the dependence of the integrality of quantumnumbers on the dimensions of configurational space indelibly stampedthe new methods as heuristic. Perhaps, the obscurity of the source ofSchrodinger's equation accounts for the meandering of the stream ofthought-a sufficient reason. What is strange, however, is the regardingof the obscurity itself as necessary and the meandering itself as final.

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