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The purpose of this experiment was to investigate the interaction of different dyes to mammalian cells and tissues, to thereby determine their effectiveness in comparison with each other.

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INTRODUCTION The purpose of this experiment was to investigate the interaction of different dyes to mammalian cells and tissues, to thereby determine their effectiveness in comparison with each other. Although the purpose of histological research is to try to study tissues in their most natural state this is impractical as most tissues look fairly similar without some kind of preparation. For this reason staining has become one of the most important procedures in the diagnosis of pathological conditions, as staining cell samples allows in-depth analysis of almost every aspect of the cell. For this experiment two of the most widely used dyes were used, those being methylene blue, a cationic stain, and haemotoxylin and eosin, a complimentary stain which is made of a cationic and a anionic mixture. The purpose being to decide which of the two stains is the most suitable for the use in analysing rat kidney tissue samples. Each dye will be used in various mixtures and pH values (as per the method) to see which dye produces the best results, i.e. the clearest and more distinguishable cells and organelles. When soaked in dye the cells and organelles change colour this is due to a process of ion exchanges. In each cell the chromagens (the staining part of the cell) exchange the cations in the tissue, such as Na+ and H+ with larger ions from the stain. ...read more.


RESULTS In the following results section the figures refer to the many attributes of the stains employed and the results achieved. Fig 1. Displays the effect on methylene blue stains when the pH level differs. This was simply achieved by using varying pHs in two different solutions. The first solution contained buffers so as to maintain the desired pH level even when the methylene blue was added. The change in pH level had a major effect on the slides as the differing solutions enabled organelles to be viewed with varying degrees of clarity. The second series were without buffers. These samples had some organelles appearing more strongly than others. Fig 2. Details the effect that time has on the H & E stain. This information shows that there is a degree of over and under staining when the stains are left for a period of time. Fig 3. This shows the effect of the solutions used to "blue" after initial staining had taken place. As can be seen the differing solutions produced images of varying clarity. Fig 4. This table shows the effect of substituting eosin for fast green FCF. This was proven to be detrimental as the very few organelles were clearly visible and the slide was extremely heavily stained. Fig 1 pH solution with buffer solution without buffer 2.6 Only the general structure of the membrane visible, little clear detail seen. ...read more.


The reason for this is most likely the result of distilled water having a neutral pH, as with the low concentration of ions there will have been little effect on the cell structure. Whereas to use a high or low pH would have potentially damaged the cell. As the results clearly showed, the use of Fast Green FCF instead of eosin in was of no use as the stains were of very poor clarity. It rarely showed all the organelles and in some cases none at all could be seen. When using the methylene blue stain it became apparent that the pH 5.6 solution (with buffers) was the best for attaining clear and precise staining. The pH value changed to 4.4 for non-buffered solutions when using the methylene blue. Out of these two different staining practices the buffered solution stain gave the clearest and most balanced staining. This enabled the identification of almost all of the structures that the staining and microscopes would allow. From the results of experiment I believe that the H & E is a much better indicator to be used when analysing the kidney tissues of a rat. The reason for this is that although the Methylene Blue did produce very clear slides. The overall clarity due to H & E's ability to colour differently cellular structures made differentiation considerably easier than when trying to decipher organelles which are all the same colour and similar looking. COMPARISON OF THE STAINING PROPERTIES OF METHYLENE BLUE WITH THOSE OF HAEMOTOXYLIN AND EOSIN JOSEPH COLLEDGE BMS 103 ...read more.

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