First Step of the Endospore Stain: Malachite green, the primary stain being applied slides of bacterial smears, heating over a water bath.

Gram, Acid-fast & Endospore Bacterial Stains - P3
First Step of the Endospore Stain: Malachite green, the primary stain being applied slides of bacterial smears, heating over a water bath.
Endospore stain showing endospores (green) and vegetative cells (red) 1000xTM.
Endospore Staining
This stain is designed to distinguish vegetative cells of bacteria (active, living cells) from endospores. A bacterial smear is first placed over a water bath and the stain malachite green is applied. It heats on the water bath for five minutes. Then the slide is rinsed and a pink counterstain (safranin) is applied for one minute. After the final rinse, when viewing the slide under oil immersion (@1000xTM) with a compound light microscope, vegetative cells will appear pink and endospores are stained a bluish green.

Endospore stain showing endospores (green) & vegetative cells (red) @ 1000xTM. Go to > More Endospore Stain Images
Sources and Resources
  • Schauer, Cynthia (2007) Lab Manual to Microbiology for the Health Sciences, Kalamazoo Valley Community College.
  • Bauman, R. (2014) Microbiology with Diseases by Taxonomy 4th ed., Pearson Benjamin Cummings.

What the Endospore Stain Reveals about Bacteria
The endospore stain is used to identify bacteria that can produce endospores—small, dormant structures akin to “bacteria seeds.” 

Forming endospores is very advantageous to the bacteria that can perform this nifty trick, such as the genera Bacilli and 

Endospores allow for survival under difficult environmental circumstances, including desiccation, starvation, high heat, exposure to chemicals and radiation.
​Page last updated 3/2016

Microbiology Lab Videos

 How to Do an 
Endospore Stain

 Also see 
Prepare a Bacterial Smear

PAGE 3                                                            <  Back to Page 1
End of Article
Learn about Gram Stain
<  Back to Page 1
Diagram of bacterial controls used for Gram, Acid-fast and Endospore differential staining.
Diagram on white board of bacterial controls used for Gram, Acid-fast-and Endospore differential staining of bacterial smears. Positive control on left, negative control on right, unknown bacteria in center.
Virtual Microbiology

The Virtual Microbiology Classroom provides a wide range of FREE educational resources including PowerPoint Lectures, Study Guides, Review Questions and Practice Test Questions.
Prokaryotic Cell, Mariana Ruiz
About Us
Back to Page 1 Differential Stains
from the free STEM 
education site 
Science Prof Online

Photographic guides to differential stains 
now available!
1. Gram
2. Acid-fast
3. Endospore

Bacterial Smears of Gram, Acid-fast & Endospore Differential Stains
This article has 
3 pages: 
1  . 2 .  3