This document provides information about gram staining, including: 1) Gram staining is a method developed by Hans Christian Gram that classifies bacteria as either Gram-positive or Gram-negative based on their ability to retain crystal violet dye. Gram-positive bacteria retain the purple crystal violet dye after alcohol washing, while Gram-negative bacteria appear red. 2) The document outlines the principle of gram staining, noting that basic dyes like crystal violet bind to the slightly negatively charged bacterial cells, while acidic dyes stain the background. 3) The requirements and step-by-step method for performing a gram stain are provided, including heat fixing a bacterial sample on a slide, applying crystal violet and

1. College of Medicine
Al-Imam Mohammad Bin Saud
Islamic University
First year
2013 – 2014
1
By : Asem Shadid – 435032467
Gram Staining
Method

2. • Asem Mustafa Shadid
• First year
• 435032467
N ame :
2

3. Microbiology lab report :
1- introduction (Gram Staining)
2-Requirements (Reagents)
3-Method
4-Observations
• Contents : -
3

4. Introduction :
4
• The Gram staining
• The Gram staining method, named after Hans Christian Gram.
• the Gram's method are commonly classified as Gram-positive
and Gram negative.
• Others that are not stained by crystal violet are referred to as
Gram negative, and appear red.
• A Gram stain and culture of the material from an infected site
are the most commonly performed microbiology tests used to
identify the cause of a bacterial infection.

5. 5
• After culture a Gram stain is performed to help determine the type
of bacteria present and to help determine what other tests may
need to be performed to definitively identify the cause of infection.
• Viruses cannot be detected with a Gram stain.

6. 6
Principle of staining
Stains → combine chemically with the bacterial protoplasm.
Commonly used stains are salts:
 Basic dyes: colored cation + colorless anion
e.g. methylene blue (methylene blue chloride)
MB+ + Cl-
 Acidic dyes: colored anion + colorless cation
e.g. eosin ( Na+ + eosin-).
Bacterial cells are slightly negatively charged ( rich in
nucleic acids bearing negative charges as phosphate groups)
→ combine with positively charged basic dyes
Acidic dyes do not stain the bacterial cell → can stain the
background material with a contrasting color.

7. 7
Requirements (Reagents)
• 1. Gather all equipment and supplies
– Gram stain
– Microscope slides
– Heat source
– Sterile swab
– Culture or specimen
– Microscope
– Label the slide with the patient’s initials or the specimen number.
2. Choose an isolated colony
off of the agar plate and
obtain bacteria with a sterile
swab.
3. Place the swab on the
microscope slide and spread the
colonies in a circular motion.
4. Heat fix the microorganisms

8. 8
Purple color
Orange color
Purple color
Method

9. 9
Purple color
White color
Purple color
White color
Purple color
White color

10. 10
Purple color Pink color
Pink color
Purple color White color

11. 11
- Blot the slide and read with the oil
immersion lens of the microscope. Look
for Gram-positive and Gram-negative
bacteria.
Pink color
White color
Purple color
Purple color
Purple color
Purple color
Purple color
Purple color
OVERVIEW

12. 12

13. Designed By : Asem Shadid – 435032467 13