Pretend you are a teacher preparing a quiz for the text I will provide. 
            Each quiz question should follow the format below. 
            Ensure that you carefully recheck the questions and answers for accuracy. 
            The output should be a ready-to-use list of dictionaries for the Python eval function:

            [

                    "question": "WRITE THE QUESTION HERE",
                    "options": ["A) OPTION_1", "B) OPTION_2", "C) OPTION_3", "D) OPTION_4"],
                    "answer": INDEX_OF_CORRECT_ANSWER_IN_OPTIONS_LIST,
                    "explanation": "EXPLAIN WHY THE ANSWER IS CORRECT"

            ]

            Text:
            positive bacteria: If they retain the primary dye complex of crystal violet 
and iodine (purple or blue color). 
 B-Gram negative bacteria: If the decolorization occurs and bacterial cells accept 
the counter stain. (red color)
 
Procedure of  gram  stain: 
• Add primary stain /crystal violet stain (purple or violet color)  on fixed smear for 
1-2 minute. 
 • Wash with water  
• Add iodine solution (brown color ) for 1 minute. 
 •Wash with water  
• Add decolorizer 
 • (acetone alcohol) (colorless) Wash with water for 15 second. 
 • Add counter stain/ safranin stain (red or pink color) for 1-2 minute. 
 •Wash with water  
• Dry the slide with air. 
 • Examine the slide with microscope by using 4x ,10x, 40x, and 100x . 
 
 
 
 
The results: 
 • Gram positive bacteria appear with purple or violet color due to crystal violet and 
iodine complex  
• Gram negative bacteria appear with red or pink color due to safranin stain. 
 
 
 Ziehl- neelsen stain: 
      This is special stain used to stain Mycobacterium spp. cell wall is thick and rich   
with lipids, wax, and have ability to resist acid so it is called (Acid fast bacilli). 
 
 
 
Mechanism of Ziehl-neelsen staining method: 
• The mycolic acid presents a barrier to dye entry. 
 • Thus it is partly overcoming by adding basic fuchsine with heating. 
• The AFB (Acid Fast Bacteria) resist the decolorizing and not losing the dye. 
 
Procedures: 
 • Add carbol fuchsine to the fixed preparation, allow to  
act for 5 min. heat intermittently without boiling the stain.  
•Wash with water  
• Add 20% H2So4 for 1 min.      
•Wash with water  
• Acid alcohol solution. 
 • Apply 95% alcohol for 2 min. 
• Wash with water  
• Wash with water, and apply methylene blue counter stain for 15 seconds air dry 
and see under microscope: 
 1. AFB:  Red bacilli  
2. Background: Blue 
 
 
 
Ziehl–Neelsen stain (acid-fast stain)  
• Differential staining 
• In this type some bacteria resist decolorization by both acid and alcohol and hence 
they are referred as acid fast organisms.  
• This staining technique divides bacteria into two groups namely acid-fast and non-
acid-fast.  
• Mycobacterium tuberculosis is the most important of this group, as it is 
responsible for the disease called tuberculosis (TB) along with some others of 
this genus Tuberculosis disease can be diagnosed in lab by the following : 
1-Taking three successive morning sputum sample from patient for three following 
days. 
 2- Making a fixed smear from the sputum sample and stained with Ziehl- Neelson 
staining method   
3- If the results were positive (see the red bacilli in smear with blue background), 
the patient must be infected with tuberculosis disease. 
 


            Quiz: