Bitera$ &anica Buena'entura$ (elsie Cantre$ (od)in Department of Biological Sciences, College of Science University of Santo Tomas, Espana Manila 1051
Castillo$ *amila
&ate !ubmitted+ ,ctober $ 2016
Introduction
Procedure
10
µ
L o. /aramecium s susension culture )ere laced in t"e center o. a glass slide
and a co'er sli )as lo)ered gently abo'e a'oiding traed bubbles % microscoe )as .ocused using t"e scanner 3 and its lig"t source )as ad4usted to reduce glare T"e ob4ecti'es )ere s"i.ted to L/, to see 'ery motile cells %.ter)ards$ a stri o. .ilter aer )as inserted on one side o. t"e co'er sli to dra) out some excess li#uid T"e sides o. t"e co'er sli )ere coated )it" etroleum etroleum 4elly using a toot"ic5 toot"ic5 to re'ent t"e slide .rom drying out T"e ob4ecti'e )ere t"en s)itc"ed to /, to identi.y t"e cell structures structures t"at can be seen in t"e unstained cells %ll o. t"e stes )ere reeated to add Lugols iodine$ met"yl green and nigrosine but at t"e ste )"ere )"ile t"e )ater )as dra)n out .rom one side )it" a .ilter aer sli$ a dro o. t"e c"osen stain stain )as laced on t"e ot"er side
Results and Discussion
Cell staining is a basic rocess in studying microscoic organisms 8n t"is rocess$ seci.ic arts o. t"e cells and ot"er comonents o. t"e cells are being s"o)n deending on t"e stain used and t"e tyes o. cell staining tec"ni#ue t"at )as emloyed !tains "a'e di..erent mec"anisms in mar5ing t"e cell 8t in'ol'es seci.icity and ermeability /aram /arameci ecium um s s is a unic unicel ellu lula larr cili ciliat ated ed rot roto9 o9oan oan$$ )"ic )"ic" " is comm commonl only y used used as reresentati'e o. ciliates in s"o)ing t"e mo'ement and mec"anism o. cilia T"is microorganism
Cell and Molecular Biology Laboratory First Term 2016-2017 can be .ound in almost all o. t"e bodies o. )ater esecially in .res")ater T"ey are tyically elongated and o'oid microorganisms T"eir bodies are co'ered )it" cilia$ )"ic" also acts as t"e locomotors organ o. t"e microorganism :nder L/,$ unstained /aramecium s$ can be obser'ed as .ast dot-li5e structures mo'ing around :nder /,$ t"e inside o. an unstained aramecium is comosed o. a 4elly-li5e .luid called rotolasm Bits o. .ood 'acuoles and ot"er materials .loat around in t"e rotolasm T"e outline o. t"e cell membrane can also be identi.ied
8mage 10 :nstained /aramecium under L/,
8mage20 :nstained /aramecium under /,
!e'eral dyes are being used in 'i'o imortant cell comonents and arts ,ne o. t"e stains t"at are being used in 'i'o is t"e Lugols iodine Lugols iodine or Lugols solution is commonly used in laboratory in detecting t"e resence o. starc" and as an antisetic o. emergency drin5ing )ater /etru99i$ et al$ 20103 T"e stain is also being used in oral cancer detection 8t is comosed o. iodine and otassium iodide dissol'ed in distilled )ater 8n t"e resence o. amylose$ a solution colors )it" dee blue ;"ile in t"e resence o. amyloectin$ it gi'es a urle color$ )"ic" is t"e original color o. Lugols iodine 8n t"e resence o. glycogen$ )"ic" is resent in t"e animal cell as storage o. glucose$ it gi'es a reddis" bro)n color T"e Lugols solution is being ta5en u by t"e glycogen .ound in t"e cell Bas.ord$ etal$201<3 T"e
Cell and Molecular Biology Laboratory First Term 2016-2017 iodine .orms comlexes )it" t"e olysacc"aride in t"e cell in t"e rocess o. comlexation reaction T"roug" t"is stain$ t"e cilia$ )"ic" are t"e locomotory organelle o. t"e microorganism$ became 'isible and aear as reddis" bro)n color T"is is due to t"e resence o. glycogen t"at ser'es as energy storage$ ro'iding t"e re#uired energy .or t"e locomotory organelle
m8
8mage <0 /aramecium cells stained )it" Lugols iodine under /,
Met"yl green is a basic or cationic dye t"at is being used in staining c"romatin &=% :memura$ et al$ 200<3 T"is stain is commonly used )it" yronin in di..erential staining o. &=% and >=% T"e "os"ate grou o. t"e &=% is negati'ely c"arged &ue to t"e cationic nature o. t"e met"yl green due to t"e resence o. t"e ositi'e c"arge o. t"e amino grou in t"e structure$ it reacts )it" t"e anionic "os"ate grou o. t"e &=% .orming a blue green color 8n t"e /aramecium s$ t"e nucleus became 'isible as a blue green structure bounded by nuclear membrane rior to t"e staining o. met"yl green T"is is due to t"e resence o. t"e &=% in t"e nucleus o. t"e microorganism
Cell and Molecular Biology Laboratory First Term 2016-2017
8mage 0 /aramecium stained )it" met"yl green under /,
Met"yl green and Lugols iodine emloys ositi'e staining 8n ositi'e staining$ t"e cells )ere t"e ones being stained 8n contrast$ nigrosin relie.$ )"ic" emloys negati'e staining$ t"e bac5ground )as t"e one being stained T"is )ay$ t"e secimen or t"e cells aear against t"e dar5 bac5ground =igrosin is a mixture o. blac5 dyes suc" as nitroben9ens$ aniline and aniline "ydroc"loride$ "eating toget"er )it" coer or iron catalyst /resser$ et al$ 20123 T"is blac5 dye .orms a blac5 bac5ground$ )"ic" enable to obser'e t"e resence o. t"e transarent /aramecium s T"us$ it re'eals t"e sur.ace details o. t"e microorganism
Cell and Molecular Biology Laboratory First Term 2016-2017
8mage ?0 /aramecium cells stained )it" ni grosin under /,
