Cell wall staining by Chances method

Please watch the video given below

Introduction

  • Cell wall staining technique is a special staining technique in which we especially stain the cell wall of the bacterial cell.
  • As we know cell wall is the outer most rigid covering of cell and depending upon the structure of cell wall bacterial cell are classified in Gram positive and Gram negative cells.
  • Staining of bacterial cell wall helps us in a demonstration of the cell wall.
  • There are various staining techniques used to stain cell wall like Chances method, Ringer’s method and Dyar’s method.

Aim

  • To stain the bacterial cell wall by Chances method.

Requirement

  1. A Clean grease free slide.
  2. Fresh cell suspension.
  3. 0.5  %  New fuchsin solution.
  4. 0.5  %  Congo red solution.

Procedure

  • A clean grease free slide is taken.
  • The smear is made on a slide by using a sterile wire loop.
  • The slide is air dried and not heat fixed.
  • After air-drying the slide is flooded with 0.5  %  New fuchsin solution and allowed to react for 3 mins.
  • After 3 mins excess stain is drained out and the slide is flooded with 0.5  %  Congo red solution and kept for 4 mins.
  • Further, the slide is gently washed with water.
  • Air dried and observed under oil immersion lens.

Flowchart of procedure of cell wall staining by Chances method

 

Cell wall staining procedure

Cell wall staining procedure

Mechanism

  • First, the smear is made on the grease free slide here there is no heat fixation step because heat fixation alters the structure of the cell wall.So here the slide is air dried and stained.
  • As we know cell wall and cytoplasm are acidic in nature where cell wall is more acidic than cytoplasm.
  • So when we apply the first stain that is 0.5 % New fuchsin which is the basic stain it stains the cytoplasm as well as the cell wall.
  • But here we want to stain the cell wall so the smear is treated with the second stain that is 0.5 % congo red solution.
  • As we know congo red is a selective decolourising agent it selectively decolourises the less acidic portion that is cytoplasm.
  • Hence cell wall remains stained by 0.5 % New fuchsin stain.

Observation

  • Under oil immersion objective we can observe pink coloured cell wall and colourless cytoplasm.

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