His to Chemical and Biochemical Studies on Leukocyte Alkaline

8/2/2019 His to Chemical and Biochemical Studies on Leukocyte Alkaline

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1955 10: 1120-1131

E. WILTSHAW and W. C. MOLONEYPhosphatase ActivityHistochemical and Biochemical Studies on Leukocyte Alkaline

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H is to chem ica l and B ioch em ica l S tud ies on L eu kocyteA lka line P h osph atase A ctiv ity

By E. W ILTSHAW AND W . C . MOLON EYT H E D EVELO PMEN T of h is tochem ical m ethod s in the past tem i yea rs hasresu l ted in a m ore fund am en ta l and d ynam ic approach to m iiany cy to log ic

p rob lem s, p a rticu la rly in the field o f hem a to lo gy . N lo re rece titly , eff icien t n ie th -od s fo r sepa ra ting leuk ocy te s from the pe rip he ra l b lo od h av e been ava ilab le,p erm itting b io ch em ica l m easu rem en ts o f enzym e and o the r ce llu la r com istitu en ts .The u se of bo th h is to ch em ica l and b iochem ical m ethods hav e dem uom is tra tedm arked d ifferen ces in enzym e activ ity in ce lls tha t appear m o rpho log ica lly sins i-la r. O f even grea te r im po rtan ce , th ese procedu res m ay b e em p lo yed ins the eva lu -a tion o f ph ysio lo g ic am id pa tho lo g ic in flu ences on ce rtain leuk ocy tic ac tiv itie sand fu nc tions . In itia l in vestiga tion s in to son ic a spec ts o f these p rob lem iss atep resen ted in th is repo rt.

ME THOD SHistochemica l: G om ori s c o i)a lt meth o d fo r alk alin e pho sph ata se , w i th n iod i fications,

w as u sed th rou ghou t. Sm ears o r im prin ts w ere tak en on co ver s lips o r s lides , a ir-d ried , andth en fix ed in 95 per cen t e thy l a lcoho l. C e llo id in w aso t used on the prepa rat ion s sin ce itin te rfe red w ith th e pen etra tio n o f th e cou nte rsta in . T he spec im ens w ere th en inm cub atedfor tw o hours at 3 7 . in a barb ital bu ffe red m edium con tain in g B eta -g ly cerophospha te ,ca lc ium ch lo rid e and m agn esium su lph ate at a pH o f 9 .2 . N ex t the cov er s lips w ere tho r-ough ly w ashed in d ilu te calc ium ch lo ride so lu tion and imm ersedn 1 p er cen t cob alt n i tra tefor f ive m inutes. T r ia ls w ith va riou s d ilu tio ns of th e cob alt so lu tio n fm on s 0 .1 per cen t to2 p er ce n t w er e m ad e a n d w h ile good r esu lts w e re o b ta in ed w ith as litt le as 0 .1 p e r c en t,0 .5-1 p er c en t so lu tions w ere fo und to b e op tim al for conssisten t resu lts ra th er th an the 2per cen t sugges ted by G om ori. T h is a lte ra tion redu cedrtifac ts an d d iffu s ion i d iff icu ltie s a swell a s ex cessiv e coba lt p rec ip ita tions on the sp ec im ens . F o llow ing co ba lt imm ersio n , thesm ears w ere w ash ed in tap w ate r an d placed in d ilu te am m onium su iph ide forfu rth e r fivem inu tes a fte r w hich they w ere w ashed once m ore , d ried , coun te rs ta ined w ith W rig h t o rW righ t-G iem sa and m oun ted . T his techn ic gave good cy to lo g ic and histochensic alv isu -aliz atio n . A s show n in P late 1 (fig .), the b lack prec ip ita te o f co ba lt su lphsid ew as depos ite din the cy top lasm at th e site of a lka line ph ospha tase ac tiv ity . T he in te nsi ty o f the reac tioncou ld be rou gh ly qu an tita ted on a plus/m in us to 4 plu s ba sis.

Biochemica l: The m ethod fo r determ in ing alk aline ph ospha tase o n sepa rate d w hite b loo dcel ls wa s th at used b y V alen tine an d B eck2 w ith out m odif ica tion . H ow ever th e n ieth od o fcell s ep ara tion w as tha t su ggested by E .le in .3 T his techn ic invo lved the p la cin g o f 0 .3cc. o f ED TA (sequ es trene) and 0 .5 cc . o f esp ecia lly selec ted dex tran in to th e barrel o f asyr ing e for ev ery 10c. o f w ho le b loo d w ithdraw n . T he sy ringe w as then in iverted at least5 tim es to in su re adeq ua te m ix in g andh en c lam ped no zzle up perm o st to allow for sep a-rat ion . A fte r stand ing fo r0 m inu te s, the p la sm a con ta in in g w hite cellsnd p late le ts w a .sfo rced ou t o f th e sy ring e in to a test tub e. T h is m e th od p ro v idedric h w h ite b lo od ce llsuspension usua l ly con ta in in g le ss th an 1 pe r cen t red ce lls . A fte r washin ig th e cel ls in sa lin e

F rom th e H em ato log y Labora to ry , I &II M ed ical S erv ices (T uftsi, B os ton i C ityHosp i ta l .

A ssisted b y a gran t from th e Am erican C ance r S oc iety , M assachu setts Divisions.W e w ould like to acknow ledge th e techn ica l h elp g iven by M iss L . F h iege ln iann th is

study .S ubm itted M a rch 8 , 195 5;accep ted fo r pub lica tio n Ju ly 11 , 19 55 .

1120

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8 6

F : . W IL T SH AW AND W . C . M O LO NE Y 1121

N OCA S E S

N Op . BL

N O .BONEA A R R O V

N O .Pt .EEN

N Oo E

NORMAL 1 4 3 1

ACUTE LEUKEMIA 9 1 2 9 I IC HRON IC MYELOGENOU S

LEUKEMIA LYMP HO CYTIC7

1 03 8

2 7

2

3

I

I 2

MYELO ID METAP LAS IA 4 8 I

P O LYCYTHEMIA VERA I 2INFECTION(PYOGENIC)

MALIGNANCY

LEUKEMO ID C IR RHO S ISREACT IONS IN FEC T. MONO .

TUBERCULOS ISOTHERINFECTIONS

MULTIP LE MYELOMA

5 2 1 4

6 5 2 I1 2 1 3 83 57 6 I5 3 3

9 1 5 1 0 -

HODGKIN S D IS EAS EAN D

LYMPHOMATA1 2 2 5 4

P ERN IC IO US AN EMIAAN D

FOLIO AC ID DE F IC IENCY7 8 6

MIS CELLANEOU S COND ITIO NS 3 9 3 7 2 7 4 2TO TAL NUMBER S 1 5 0 2 5 6 8 0

TABLE I S umm a ry o f th e c a s e s a nd m a te ria l s tud ie d fo rlk alin e P h os ph at o s eb y t h e h is to c h e m ic a l t e c h n Iq u e

and centrifuging tw ice, only mininial platelet contaminations was found in the last suspen.sioI l, an iml)ortanst factor in oh)taining leukocytes free fronu agghiutination.

MATER IALSAs shown ins table 1, 150 patiensts were studied and divided into various disease groups.

The niaterials consisted of peripheral blood and hone marrow smears and imprints or smearsfronts spleens and lymph nodes. Ins all, 350 histochemical procedures were carried out on thesespecinisenis and inmany instances biocisemical determinations were made on separatedleitkocytes of these patients. Thse major part of this investigationwa s concerned withletmkensia and allied disorders; however, in a( ldition normal controls and variotms other dis-



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1122 LEUKOCYTE ALKALINE PHOSPHATA SE ACTIV ITY

orders w ere studied. Inc luded in this m isce lhanseous g roup w ere hemolytic , hi pochronssic andaplastic anem ias, purptmras and lupus ery thematosus. A lthough sonic intere sting observ a-tionis w ere made , no spec ific fe atures of alkaline pisosphataseactiv ity w ere found in thisheterogeneous g r o u p . While 7 cases o f megaloblastic anem iawere inve stig ated, intercurrentinfec tions or o ther complicating factors nuade i t inspossible to ins terpret thie hndinsg s.Gener a l O bser va tio ns

Wachstc iui and others4 state thatn the peripheral blo od omilyhe seguisemited

100- NORMAL(AVERAGE O F IICASES )

pos itiv ely sta ine d ce lls80 - EJ % negatively stained cells

(I)

Li.. 23 .0 7 mgs. of P libera ted in I hr.60 - by 10 W BC sz

m0z(I)I0. . 20 7

/0

__4 t t 2t 3t 4 tI -

90 - /

70- PYOGEN IC IN FEC T IONo / 161 .6 mgs. of P libe ra ted in Ihr.w by lO b WBCs

I50-

w0w

//lO _ ______

shows the ave rage distribution of sta ined ce lls and theCHART I mgs of P re leased in Ihr by 10 10 W BC sn normals

and in cases of pyogenic infection



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C HR O N IC MY E LO G E N O U S LE U K E MIA

% p o s itiv e ly s ta in e d c ells

E J % n e g a t iv e ly s ta in e d c e lls6 6m g s . o f P lib e ra te d in lh r.b y IO W BC s

4 0

2 0

MYE LO ID ME TAP LAS IA

II6m g s .o f P lib e ra te d in h r.b y lO # {1 76 }BCS

E. wILrsuuAw AND W . C . M OLONEY 1123

ama! I ntuit! forums mieutrophils comita iu i alkaline phosphatase. Ami adequate counter-stain 101(1 a shout iuictmbatioms period unade it. apparemit imidecd that this emizymise is(lemonsi rable univ ins the cvtoplasmii of these peripheralblood! cells amid that it(lOes mu t t appear iui the nucleus. I t has also beemi said that Isistocheunical methodsfor alkali me phosphat ase were ummsat isfactory iiibomie marrow preparat iouis, butthis has not beeui ottt experiemice, if thimi mnarrow smearsare umsed!. Except for anoccasiouial musetaunvelocyte amid for a rare histiocytic- like (dl (see Plate 2)fouumd iui certaiui disease states, the alkalimme phosphatase was couif imme(! to theadult amid hauid mieutrophil i im thelmie miiarr()w . This was true also iii the spleemi

zw0

wa-

s h o w s th e a v e ra g e di s t r i bu t i on o f s ta in e d c e lls a n d th eC HAR T 2 m g s o f P re le a s e d in lh r. b y 0 1 0 W B C S in c hro nic

m ye lo g e n o u s le u ke m ia a n d m ye lo id m e ta p la s ia



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1124 Ll:UK0CYTE ALKALIN E PIIO5P1IA TA SI: A (rmv LIV

PL A T E 1 : F t c , . 1 -Peripheral blo od smear front a case of chronic nivelogenstitnss t a i n e d f o r a l k a l i n e p h o s p h a t a s e b y t l i eonsori I e c h n i r .

amid! lymph miode miiaterial aswe l l as im i use ce lls obstaimied Irom ii various bodyfluids.Vorni al Per ipheral Blood

I mi mornssal subjects t.he msuajo rity of po lym isouphiom ismeleam leu k scytes shsW d( I lit)s taim i fo r alkalimie pho sphatase. A pproximately 20-4 () u cent (It ce lls meV eale ( la plims nusim ius to 2p l u s s t a i mi i u mg i mi t m usity . B iochiensical deteruiii umat( uis gave au



7/13

Pn.TF: 1 : Fir. 2.-Peripheral 1)100(1 sm isear front a case of ni lo id nie taplas ia stained foralkaline phosphat ase by te Gonsori teclinit

1 :. W ILTS IIA \V . N1 ) w . c. MOLOXEY 1 l2

average r e s u l t o f 2 1 . 9 mu g . of P liberated per hour by0 # { 1 7 6 }euko cy tes, w ith araumge of lS -2 1 i mug .

InfeelionJut all cases o f pyogemiic im ifec tioms the leukocyte s show ed greatly increased!

alkalim ie phosphiatase ac tiv ity . In omie case o f W eils (lisease amid iii several caseso f pim itisom iary tuberculosis , msiarked e levatiom i of em izyrne activ ity w as also oh-



8/13

1126 LEUKOCY TE AL KAL I NE PHOS PHAT AS E ACT I V I T YTAB L E 2 Showing the distrbution and tota l content of Athdine P h o s p h o t a s e in cases of multiple mye loma o,idchroesc l y m p h a t i c le ukem ia compared w ith th a t in normals ond chronic m y e lo g e n o u s le u k e m ia c a s e s

D IAG N O S IS NOC A S E S

AV NOP O L Y S

PE R CM MAV .NE G

CE L L SAV P 05

C E L L SA VE RA GE D IS TR IB UT IO N

- - -

+ 2 + 3 + 4+L EUKOCYT E

AL KP H O S t o

POLYMORPHAL KP H O S 4

N O RM A L 8 4 ,8 0 5 6 4 4 0 3 5 6 0 1 2 0 1 8 0 5 6 2 8 9 6 34

CHRON ICLYMPHAT ICL EUKEM IA

3 4 . 2 4 1 3 5 1 0 6 4 9 0 l2 0 2 8 3 1 8 0 660 17 3 7 1 1 5

8 4 .5 3 ) 2 8 3 5 7 1 6 5 1 4 I 2 3 4 10 7 8 7 5 1 4 7 4 3 1 0 6 4

CHRON ICMY E L O G E N O U SLEUKEMIA

3 5 7 .3 0 7 9 8 7 0 I 3 0 I 0 0 3 79 0 1 4

o rA l h a l i n e P ho sp ha to se o ctios ty w ip re ss ed os mgs o f P lib eis te d p ee hour b y 0 0 leukOcyles5A l h o l i n e P ho sp ho to se a ctivity e x p r e s se d a s mgs o t P l ib e r a te d p e r tr e e b y lO n e u t in # {2 1 6 }s ilia ymorphonucleor leukocy slo&lt B bond forms)

served. It. w as interesting that incases o f im ifec tious momiomm imeleos is , the alkalinep hosp h ata se ac t iv it y w as tseghigible it i the uteutrophils o fthe peripheral blood.Lenkernoid Reactions

X eutrophil leukenso id reactions are seem i its a varie ty of (lim lical (hsordlers o therthais in response to pyogenic im ifec tiout. lu-i this series 4 cases o f myelo id us ie ta-phas ia, 6 cases o f carcinoma am id 12 cases o f c irrho sis o f the liver w ere studied( table I). Other observers have show ms that thse po lymorphsomiuclear leukocytesunder these conditio tis have high ce llular alkaline pho sphat.ase activ ity by histo -che tiiical and biochemical measurenseuits . S im ice these diso rderssiav sius iulatechromiic nsye logenous leukemia c lose ly thsese (le term isim iatiom is are of puattitalvalue as aus aid in differeustial diagnos is.Leukemia

It is s e ll kmsown that iii acute leukemis ia the imsiniature mseutrophilic leukocytesare devo id of alkalimse phosphatase . Ims chrousic msiye logenous leukemia this cm i-zyme cammot be demonstrated except as a faust plus /m isim ius stain iui am sccasiomia lce ll. B io chem ical readings w ere also veryow w ith an average of 7 .8 mg ofliberated per hour by 1O#{ 1 76}hite blood ce lls .

Charts 1 atid 2 and Plate 1 (figs. 1 and 2) deinomsstrate c learly that leukocytesthat are morpholog ically s im ilar may contaiui niarkedly diss im is ilar alkalim ie phos-phatase activ ity . The high comitem it o f this euszymell myelo id tise taplasia aisdpyogenic infec tio n is in marked contrast to the low miormal ce llalue amid thealmost mseg lig ible alkaline phosphatase activ ity foumsd in the leukocytes o f chsrous icmssye log enous leukemia.

The biochemical measurememit abuse may be mis leadiusg as an im idex of umsitm ieutrophil alkaline phosphatase activ ity . This became apparemst ims the 9 nsul-tiple myeloma and 10 chronic lymphatic leukensia cases s tudied ins this series.While alkaline phosphatase activ ity appeared normal by the biochemical tech-nic, the his to chensical method show ed intem ise staining of almost every adultiseutrophil. B y (orre latim ig the biochemical readings to0 10 s eg us ie mi te d amid band



9/13

E. WILTSHAW AND \V . C . MOLONEY

PLATE 2 .-H istiocytic -like ce ll in hone ns arrow show ing pos itiv e stainning fun alkalineplto sihat ase ac tiv ity in cy toplasm.

formss neutrophils it because obv ious that im i these dhseases unite ll alkalim me phsos-phatase activ ity w as g reatly e levated (table 2 ) .

V alemstim ie6 has showmi by biochetsiical methods that. the high lettkocy te alka-lim se phosphatase ac tiv ity im s pyog enic insfec tiouss dim iuiishes as t.he clinicalo m i -d!ition improv es . These fiusding s w ereconf irmed iui our studlies by both biochemi-cal and his tochemical methods amid are s h o w n graphically in Chart 3 .

1 1 2 7



10/13

1128 L E U K O C Y T E A L K A L IN E PH O S PH A T A S E A C T IV I T Y

ou,

0o.-Jo

0 > -L m 5Q -o .125 H

C i- P E N IC ILLIN TH E R A P Y IC ))a -zLI- 25 - / num ber positive cells /cm m . 20 : : : Jnumber nega tive ce lls/cmm .Co(1)1 0 -5 . ,01 - /

z 0 3 8 13 DAYS show s a fall of A lkaline P hosphatase A ctivityz C H A R T 3 concom itant w ith clinical im provem ent

Experimental ResultsTise fumiction o f alkalim ie phosphatase iii the n eu t r op h il is u m ik u iow n . It h a s

bee ts postttlated by V aleuttim ie and o thers that this enzymeay p l a y a r o l e i mithe reac tion to infec tion or o t h er st r ess p h en om en a . A nother po ssibility is that.this em izymsse takes part im i the intrinsic nsetabo lic activ ity of the (e l!, such asg lycog emies is or nuc le ic acid synthesis. It hasc curred to us that the neutrophilma y be a vehic le for the tratisport o f this enzynse frons the bone ssarrow to othersite s . There fore , attempts w ere madeo study the re lative concentration of alka-l im ie p ho s ph ata se i n the leuko cy tes o f peripheral blood and bone marrow whenthey w ere dlraw n s imultaneously . A lthough erro rs such as dilutiom i o f bone mar-row aspitates caniio t be exc luded, itppears, so far, that his t.o chem siically andbio chemis ically there is ito co tssis te ts t difference betw een these nnaterials (seetable 3 ) . However, further studies are be ing undertaken to te st this hypothes is .



11/13

TABLE 3 A compar i s on o f A lk alin e Ph o h a t a s e a c t iv it y o f b lo o d a nd bo n e m o rrow m a ro us c o nd itio ns

E . W LTSHAW AND W C. MOLONEY 1 1 2 9

AGNO S 1 S CAS ENO

S O UR C E A V E R A G E DISTRIBUTSON- - - - -

+ 2 + 3 P 4tV.

POS ITIVECELLS

V.NEGATiVEC E L L S

LEUKOCYTEA L KPHO S . *

POLYMORPH .A LK

P H O S

N O R M A L I P BB M8 I I I

--6 I

- -3 6 6 4 3 4 . 8 49

14 30 2I I -66 34 3 4 . I 81MUTIPLEMYELOMA 2

P BB M

3-

25 24 7 5 9 4 1 6 1 0 9 38 19 I 2 15 26 8 ) 1 9 33 0 62

RETICULUMC E L L S ARCOMA

PBBM

3 3 9 413

I 139 2 2 2 6

9 680

42 0

OO5 3 0

73 6 5

HYPOCHROMICA NEM I A

PBB M

4 2 - - - 2 6 7 4 2 3 0 281 0 5 2 27 7 3 IS O 3 1

LEUKOCYTOSISC A U S E ?

PBB M

1 3 2 1 - 8 - 4 2 5 8 2 0 0 2 24 10 6 3 23 77 50 29

5 A l k a I i o w P ho s pha ta s e o c he ey s o p ie ss e d a s w ig s o f P h b e ra t e d p e r hiat t b y t O o c y t e st A l k a l i n e P ho spha ta se a c tiu y e ste em e d a s r a g s . o f P lib e ra te d p e r h o u r b y10 neutroptt po lym orp ho nuc le ar le uko cyts s lo d u l t B bond forms)

A nsom ig other variables w hich issay effect em izynse activity is the period of im icu-batiom i of the cells in their ow ui serum . A s show n in C hart 4, w hen lem tkocvtesw ere im ictm l)ated intheir ow n serum the em szyusse activity doubled itself im tbout2 hottrs am id rose gradually to reach equilibrium betw een 4-6 hours.

D ifferem ices its alkaline phosphatase com icem stratiom s im s the leukocyte m ay hecom iditioned by (1) enzym iie activity in the serum , (2) avidity of the cell for theem izym e, or (3) intritssic control by souse nsecham iisni im s the cell itself. In thestudy of this problem the lack of a relat.iom iship betw een sem uns alkaline phos-phatase aisd leukocyte alkaline phosphatase w as confirm ed. E xperim iiem its w eret.hem i perfornsed usim ig w hite blood cell suspensiom is froni cases of m sornial sul)ject.s,of chronic m yelogem sous !eukeusiia, of a case of usiyeloic! urietaplasia am id of pyo-gem tic infectiotis. The cells w ere im scubated for om ie hour at7 C . im i sahim ie andvarious sera (see table 4). Follow im ig inicubatiom i alkaline phosphatase w as dieter-ussiused l)iochem iiically in the usual m am inerm id! new blanks w ere rum ito (orrectfor the possibility of aus im icreasein the intracellular inorganic phosphorus. Inall cases the cell alkalim se phosphatase rose during inicubat.ion but this w as im i(le-pem idem it of the seruni or salim ie m etistrum . These fiusdings suggest that the cellalkalim se phosphatase is riot. regulated! ins vitro by any factor im s the serumutis im itrim ssic to the leukocyte itself.

Su.t R Y(1) lus a large series ofnsdividuals alkalitse phosphatase activity im i the leim ko-

cyte sas studied! by biochensical am id histochem isical m ethods.(2) The value of com bim ied! hist.ocheniical am id biochem ical m easurem ents is

dem onstrated in this report.(3 ) In chrotsic niyelogenous leukem ia the m ieutrophil com staim s very little alka-

lute phosphatase activity w hile inm yeloid m etaplasia am id pyogetiic iusfectiouisussorphologically sim ilar cells exhibit a great increase im i this etszynse.

(4) V arious factors w hich m ight influence alkalim ie phosphatase activity im i



12/13

INFECTIO N I

INFECTION 2? MYELOID 3

M ETAPLAS IA

4N O RMA L 5

6

HOURS O FINCUBATION

1130 L E UK O (Y T E A L K A L IN E PH O S PH A T A SE A C T IV I T Y

14 0

l3020

110 -100-

90 -

80

70 -

60 -

50-40 -

30 -

20 -

l0

I I I I I0 2 4 6 8

s ho w s th e e ffe c t o f tim e o f in c uba tio n o f WBC sCHART 4in th e ir o w n s e rum o n am o unt o f re le a s e d by01 0 WBCs

WB CSUS PENS ION

BEFOREINCUBATION

INCUBATION FOR ONE HflUR IN : S E R UMALK. P HO S.

INB . U NITS

S E R UMINORGANIC P .

INMGS .

S ALIN E C .M. S ERUM NORMAL S ERUM HIGH. Fi S E R UM

NORMAL 2 8 .8 5 1 .0 7 0 .0 6 9 .0 4 5 .0 3 .0 2 2 .4 8

C.M .L t 1 5 .2 3 3 .6 2 8 .0 3 1 .4 3 2 .6 3 .0 5 2 . 1 3

H IGH A . P . * 1 3 6 . 0 1 3 1 . 0 1 5 5 . 0 1 5 7 . 0 5 7 . 0 8 . 0 7 1 . 4 9

TABL E 4 TO S HO W THE E F F E C T O F IN C U B AT IO N O F C E LLS IN S A L INE , THE IR O W N S E R UM, A ND TH ES ERUM FROM OTHER CAS ES W ITH VARYING AMOUNTS OF C ELL ALKALINE P HO S P HATAS E .

+ CHRON I C MYELOGENOUS LEUKEMI A* HIGH C ELL CONTEN T OF ALKALINE PHO S P HATAS E (A CAS E OF P YOGENIC IN FECTION



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E . W IL T S H A W A N D W . C . M O L O N E Y 1131

l enmkocy t e s w ere im iv es tigated an sd it w as f ound that ou t incubatiom i iii sem um is o rsaliuse the ce ll alk aliuse phosphatase ac tiv ity increased . H ow ev er, th isn vitrorise w as um srelated to the sou rce o f the serum m enstrum .

S UM M A R IO IN IN T ER L IX GUAI . Ju t un gratsde serie de in d iv id uos le ac tiv itate d e phosphatase alcaliusn

leu cocy t.o s essev a stud iate per m ethodos b io- e h istoch im ic .2 . L e v alo r de con ib iu sate m esurationes h isto - e h ioch irn ic e s densom istrate im i le

presem ite repo rto .3 . In s ch ronic leu censia m y e logen e le neu trophilos ha bassis sim e activ itate de

pho sphatase alcaliu s, du ram ite que in m etap lasia m y e lo ide en in f ec tiom ies py ogenece llu las q ite es nsorph o log ican ietite af f in ex h ih i un gram sde augm eutto di e illeemszy ts sa .

4 . V ane f acto res que es poss ib ilem em ite capace a af f ic er le a c t i v i t a t e d ie pho s-phatase in leu cocy tos essev a im sv estig ate . I! es sev a constatate que iuscubatio nin sero o in so lu tiom i salim i resu lta in uts augm eisto de l activ itate ce llu lar d e phos-phatase alcalin . T am em i, iste augm etito im s v itro essev a sim selatioti a! orig im se de lun em istro seral.

R EFERENCESGOMOR I , G. : M c r o s c o p i c Hi s t o c h e mi s t r y . Un i v e r s i t y o f Ch i c a g o P r e s s , 1 9 5 2 , p .84 .

2 V A L E N T I N E , W . N . AND BECK, W . S .: B iochem ical stud ie s on leu k o cy te s. (1 ) Phosphataseactiv ity in health , leuk o cy tosis and m y elo cy tic leuk em ia. J. L ab . & C lin . M ed . 38:39, 1951 .

1 \ LE I X , E .: Personal com m unicatio n .W A C H S T E I N , M .: A lk alin e phosphatase activ ity in blood am id bo ise m arrow cells. J. L ab.

& C lint. M ed . 31: 1 , 1 9 4 6 .HAIGHT, W . F. A N D R O S S IT ER , R . J.: A c id and alk alim se ph osp isatase in w h ite ce lls: D ata

f or tise lym phocy tes and th e poly m o rphonuclear leu k o cy te s o f m an and rabbit. B lo od5: 267, 1 9 5 0 .

V A L E N T I N E , W . N ., B ECK , W . S ., FO L L ET T E , J. H ., M I L L S , H . A N D L A W R E N C E , J. S .: B io -ch em ical s tu d ies in chronic m y elo cy tic leuk em ia, po ly cy tisem ia v era and o the r id io -l)athic m y e lo pro lif e rativ e d iso rders . B lood 7 : 959 , 1952 .


His to Chemical and Biochemical Studies on Leukocyte Alkaline

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