Section cutting in Microscopy

SECTION CUTTING I N M I C R O SC O P Y

SECTIONCUTTING IN MICROSCOPY

H. F. STEEDMAN PL,D. D.partnent aJZoolagy, The Ufiv+tity,

j LNs if the stecino is nall md soft, or for z+ hom at .oo- t"-po"t,," ifit ir lurd. J. Ino r \o. OO geLd! c"D,uleohc rhesorcineou.og " 6re prpene.od Lhm ld rp rhe epsLre. Thiq is bsr done ki.h a pipenr wticl tD. be{ 6led u,irh lesin."lae rle ri! ofthe lipen at rhe bofto;;fihe carsuleod gottyili the epsulc fron rhe bonon tpwrdr soth,t ao air bublle areprcdu..Jq perhittcd ro "jck rc thr sid6. rftbe speci@ moveak) 60- $; -'ddtc .ir!e 6tted opsule gody move tleo br.L .gair, $ing lhe 6ae pitefle. 6. ser_l-be res 6y krcpiig tle ep*J., witf ie co"r oD.iD s o!@ a! 48" c for 7. To rmove the gelarin pl&e rlc €lsule jn wam water, rd brush awy the geratrns rt sotreiJ. 8. Trin the @d ot dE hJock$ dL rhe speciJnenis i! rhe njddje or a , bm. :_cuir" p.. :}i1 udd a bilocvrr nia;.cope usiDsa oie+dsed safeq r/or bladc.TIe biock n r!tu reedyror cuniis. CaitingtudA e MBlo.Is rbe tudng of !h6r blocLt L &e !@e 4 lhe curiile of Dethacrvtetes eiver on p. oe.Tbr sc.tio6 er tut ob io puc w,Fr (cluen;d ctaen, r;js). Epoxt tus ib Modif catians It my te fouad h pradise rLat wfile a blo& of .Aiddite' dts we , ihe Daterial in it my nor do so. Cledly rhis is duc ro incomltete peaerrationofthe catalyzed od acelented roiq od it uy be due to ditrermdjpaebation of the dtrc (D.rn, r9J9). He puB fotu*d the view that the isdod pfletrates the riss@b€ner rbd do6 the r6in, dd thar wirL a tornlt4 where rhe r;in aad thc hrdener m prsot in eqnal xnouts (Glaen @d claudq re5a) the rises becom poneated

Aba This S&tio$ fat theELchallMilotope

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with a nixture containingincoEcct protonios. To courtcracr thls todency le lurs lorsaid the folosing ni\tue dd tining: r. Sring 6xed tisus into absolureerheol 2. Absolute ethdol dd Alaldire jo/5o at 48' C for 6 hou . I. PEe'Araldite 72 hon6. Chmge wery:4 hours 'Araldite M r5bLl-,. 4. ,. . -r t sork ir rbr []urc-orz.: hour.,rrh frohrsir ffirre 'b48$ rre'v ' 4 Djbunl DhLhJrtr r nJ I 5. Add o 3-",1. aaelentor to'the alovc mixture ad soaLfor 2a houa ar loon 6. Pla@the slecimo in a 6esh dd &elcratcd mirture ud bake thc bloct for r 7. rr tdlding of the bakedblock sti shom ir to be on the soft side bakeit lor a f!fther ts3 dq's at 48' C. EttiliMkl The 6e of ure!-forh,ldehyde reiu s infltmtio nedia for rhe cuning of seciou betwed r 20 I is giveo jn Clapter 17. Their e s bdltration hedia for ihe dtting of cctions bctweo rcr,ooo A is possible,rhough rhey shodd be regarded s experimdtal techniqus ody. The resimde soluble waro od havea visosity which is su6ciatly low, wher Eiied vith otler reg€$ts, to tueiit qeful invesr;gario$. Uscd alooc, without $ftding agents,urea-formaldchlde resnb .ie brnde, but tiey rnay be softoed sufi.idtly to clt wcl. Thc folloiring lomula is suggestedc leing suitable. Ur 600 6oopolyethylm glyol, nolto water Urea foEuldehyde sFup DzzTo Phdol dtsrals Trinerhylol lropde

35 d. ,o nl. x5 mt ) gn. ) sn,.

Tle lolyethylde glycol should be hcatedin a bsker uril it is noltm. Then the water should be added,and the other reg&ts in rhe ord* givo. rhe seqme of rnixins shouldnor be alereA Ha/dder , Aj@onim .lJoride rr.J sm. Ura 3j gm. ware! to roo ol_ Aroonir o 88 S.c. ro !H 7.j. idda6 The sho!1dbe addedro the lsin in rle lropoitior oforc &op oftudorr to ocb nillilite ofDzzTo. fl1!s roo nl. ofLrl 600shouldlave 35 drops oflardmer. sp€cirnedrtuy be brolshr inro LT 600fron wat r. Ttre liadcne. shouldbc added bclore lhc srecifro is introduced.The resin ed hddeDersLoutdbe welt hired by

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Srcti Cuuin! b Mictos@pf

stiring, and wher rhe 6ubb16 introdwd by stidlg Lavc foeted to thc top tle spccim@shouu bc .dded, and lhe rubeplaccdin a rcFigerator ar 9-roo C. The tubc shouldle corked,hd it should!e $ned at;nterals to naLe swe tlat tle oecitrm is beirg b6irrrted. A Jow ,rift* s suggekd on p. ,J b veD $iLrble. Shouldt\e speino be kept for longer tla 2a hos lhe flrnd shouldbe .hanqed. To nake a blocl pou into " gL6, rrbe $rc D.wly dixed and @Ljlyrd rsnr-and ,ddrhe specino. Anrg- rh 'pecimeb* Lhebo,ron ofdr glas rubeod leve ou thc bencha! a rooh tmpe.atue ofa" C to hsder. Tbis tehperature is dot diridl. Any tem?datue will do, bufthe lowd lhe slow6 vil te the re'h ro h..d@. Shodd there be dy .dhtage ia iDfrltratioo dd hrdeiing at J-ro' C the tube may te ld h thc reftig@lor, dd the lesin will hardo in 3-5 &1s. At 20' C dr iesin wil hrden ovemight, but should more repid h*dming !e requned the tube nay be pur inro an oven ai eronnd 30. c. Thil wil hrde the rein in abont fou hom. The tube ued for hddding crnnot le the !s$l gelatio tule baause the water still premt in tne resin solutior will sofrei rhe gelatin. The glas iube oscdsholld havc ar intemal dianets alont r nm. lreu thd the chu& of the niaorotu. whar the raia slrinks, o it wi in d@ cou;e, the blo& sLould dt dely bto tte The block vilt get hdder day afte! day. The point at whictt it i jut right for dltiog wil dependon tu plcvailng coD&rios ofloom rdperatu's etc. sectioro should be doned o! ro w3ter conraining2% glyceiol ad 2% acetoft. I1r glycerolis ttree to prerfl 6r comple.rdryingsd ortrg oftie :ecnonLttr. Tle r.etonen to redr,e fie 'uf"cc rdioo of lLr slv.erolv?Ler. Re'in Wlnatkr r. The vjscosityof dy rdn in the liquid condition is invqsety propoltion l to its tedpdlrue. Therefo.ci{it i ihpondt ro iDnlbatea s!6ioen at a low remte..tee e largeindede i! the infitratior tine shouldbe qivo. The a.rual extdioD of libe i somefiingq hj.h shouldb- s dked our by rll ud edor, bibg sr