Cold ZN - Kinyoun Stain Pack

HS805 Instructions for use v1 09.04.2015

Product identity (name) Handling and storage

HS805AMB, HS805MB, HS805MG Store in a dry, well-ventilated place and use before Cold ZN – Kinyoun Stain Pack (synonyms: None) expiry date. CAS-No: N/A Pack Size 3 x 250ml Intended Use

Product composition IVD, for professional use only.

A. HS180 - (Kinyoun) 250ml Type (s) of material contains <1% methanol (CAS-No 67-56-1), Sputum, bronchial washings, urine, body fluids <20% ethanol (CAS-No 64-17-5), (spinal, pleural, pericardial, synovial, fluids) from ~6% phenol (CAS-No 108-95-2), ascites, blood, pus, bone and tissue biopsies. ~3% basic fuchsin (CAS-No 632-99-5). [1] Suggested Procedure

Principle: B. HS787 - ZN Decolouriser 250ml contains <5% methanol (CAS-No 67-56-1), In 1915, Kinyoun published a method that has 3% hydrochloric acid( CAS-No 231-595-7), become known as the “cold staining” method <90% ethanol (CAS-No 64-17-5) because the heating step was removed in favour of using a higher concentration of the carbol fuchsin primary stain. This procedure is used to stain Mycobacterium and Mycobacterium leprae. These

C. Counterstains - are called acid fast bacilli. Acid fast bacilli stain pink/red and, the background will appear blue i.) HS500 - Alcoholic 250ml due to methylene blue, or, green due to malachite contains <5% methanol (CAS-No 67-56-1), green. <75% ethanol (CAS-No 64-17-5). The rods stain with carbol fuschin. This dye is retained when treated with acid because of the presence of mycolic acid in their cell wall.

Handling and treatment before staining: ii) HS435- Malachite Green (ZN) 250ml iii) HS515 - Methylene Blue 250ml Smear 1 to 2 drops of specimen on the slide, and Polychrome (Loeffler) allow it to air dry. Do not make the smears too thick Tissue samples may be used. Control: Any tissue containing acid-fast organisms. For further information see SDS.

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TCS Biosciences Ltd, Botolph Claydon, Buckingham, MK18 2LR United Kingdom t: +44 (0)1296 714 222, f: +44 (0)1296 714 806, e: [email protected] www.tcsbiosciences.co.uk Cold ZN - Kinyoun Stain Pack

HS805 instructions for use v1 09.04.2015

Technique Additional Information

1. Flood the fixed smears with Carbol Fuchsin (Kinyoun) (A) HS180 and stain for 1-3 minutes The quality of the staining for acid-fast bacteria will without heating. be affected by the quality of the smear. A good 2. Wash with deionised water and decolourise with quality smear will have a thin film of the specimen ZN Decolouriser (B) HS787 until the dye no or culture, allowing individual cells to respond to the staining protocol. The waxy nature of the mycobac- longer runs off the slide. teria organisms causes them to repel water so 3. Wash with deionised water and counterstain for fluids should be added to the slide after spreading 10-30 seconds with (C) Methylene Blue HS500 the sample in a thin film over the slide. or HS515, or Malachite Green HS435. 4. Wash, gently dry and examine. When cultures have not been sufficiently mixed to break up clumps of cells, the resulting smear can be difficult to read because individual cells are not discernible. Expected Results[1]

Acid fast bacilli (straight or slightly curved rods)

- red (beaded appearance) Users must determine the suitability of the kit for

Other organisms - stain blue-green. their particular use.

Background - blue if the methylene blue counterstain is used or green if malachite green is used. Bibliography

1. Kinyoun, J. J. 1915. A note on Uhlenhuth’s method for sputum examination for tubercle bacilli. Am. J. Public Health 5:867.

European conformity according to the "IN-VITRO DIAGNOSTIC MEDICAL DEVICES DIRECTIVE 98/79/EC, ANNEX III". Manufactured by TCS Biosciences Ltd.

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IFU HS805 09.04.2015 v1