CONCLUSION
From the experiment that we have done, finally, we can conclude that Gram staining is the method of distinguishing between Gram-positive and Gram-negative bacteria. Gram positive bacteria have a thicker peptidoglycan which makes up their cell walls but have greater lipid content since these bacteria have an outer membrane. The differences in their cell wall architecture cause them to look different from one another by performing Gram staining procedure. Escherichia coli appeared reddish pink which indicate that these bacteria do not bind the crystal violet stain. Therefore, it is determined to be a Gram-negative bacteria. The decolourizer extracted the lipid from outer membrane of Gram-negative bacteria washed off the crystal violet-iodine complex. afranin then was used to counterstain Escherichia coli leaving the colour pink . Staphylococcus aureus absorbed the purple colour from crystal violet. This indicates Staphylococcus aureus is aureus is Gram-positive bacteria which have thicker peptidoglycan in their cell wall that trap the crystal violet-iodine complex more effectively, making them resistant to the decolourizer. !n this experiment, we also have provided some material to help us for reaching the aim of this experiment such as "ucker#s crystal violet, Gram iodine, $% & ethyl alcohol, safranin, and microscopic slide, staining rack and bibulous paper. "owever, before doing the experiment, we absolutely need to pay attention on the precautions. There are several procedures that we have to do in order to avoid avoid the error in this experiment, such as prepare smear from cultures of microorganisms, heat fix smears, place slides on a staining rack and so on.
REFERENCES
'. (eboffe ).*ierce +. ' .Gram tain, *. '%-'$. !n )icrobiology (ab Theory and /pplication, 0rd ed., )orton *ublishing 12., nglewood, 1o. . 1appucino G.3ames, herman 4ataline, 5th edition, *earson ducation. 0.
QUESTIONS
'. !f the iodine step were omitted what colour would expect a Gram-negative microorganisms to be6 / Gram-positive6 7 !f the iodine in the Gram#s stain was omitted, the crystal violet enters the cell will not able to form the crystal violet iodine complex. The crystal violet iodine complex molecule has a larger size than that of crystal violet molecule and hence Gram-positive bacteria are able to retain stain. !f the iodine were omitted even Gram-negative cells would get decolourized by alcohol treatment and would be seen as a Gram-negative cells. 2ther than that, they would all be the colour of your counterstain 8usually safranin9 because without the aid of Gram#s iodine, which acts as a mordant 8helps to set the dye into the organisms9. 1rystal violet would not effectively set into the organisms, and when the decolourizer is added, the crystal violet will be washed away and the counterstain will be the only stain effecting the organisms. . 1ould other dyes be substituted for crystal violet Gram stain procedure6 For safranin6 xplain your answer. 7 1rystal violet dye can be replaced with methylene blue. This is because both are basic dye that have a positive charge that able to bind to negatively charge particle, such as the teichoic acid at the bacterial cell wall. Therefore, methylene blue can be used to replace crystal violet dye. )eanwhile, for safranin, it can be replaced with basic fuchsin. +oth dyes are able to stain Gram-negative bacteria
which is a pink colour but based on the information that we have searched, basic fuchsin give much brighter colour in order for the bacteria to be seen clearly. 0. :hat role does the mordant play in the Gram stain procedure6 7 The mordant is the substance that set dyes the iodine in Gram stain fixed the crystal violet to bacterial cell wall of the Gram-positive bacteria. :hen Gram positive bacteria are treated with ethanol, the alcohol is thought to shrink the pores of thick peptidoglycan. Thus, the dye-iodine complex is retained during this short decolourization step and the bacteria remains purple. ;. :hat correlation is there between cell wall composition and Gram reaction6 !s Gram reaction correlated with sensitivity too various antibacterial agents6 7 1rystal violet dissociate into 1l- ions and 1<= which interact with negatively charged components of the stain in purple. !odide ion 8! -9 is than react with 1< =, forming 1<-! complex. /lcohol interacts with the lipid of cell membrane, causing Gram-negative cell lose its outer lipopolysaccharides membrane, exposing the peptidoglycan layer. Therefore, 1l-! and the outer membrane is washed away. "owever, in the Gram-positive cell, the alcohol dehydrated the cell, trapping the 1l-! because of the thick peptidoglycan. The Gram reaction is somehow correlated with sensitivity to various antibacterial agents. Gram-positive cells are more susceptible to mechanical breakage as the amount of peptidoglycan is small. "owever, the reaction does not ensure that the same antibacterial agent is able to destroy the two 89 different species or strain of bacteria even though they showed the same result in the Gram reaction. This is because types of action such as in>ury to the plasma membrane, inhibition of cell wall synthesis and prootein synthesis, does not only depend on the Gram reaction.