This document describes several dermatological examination techniques including KOH examination, Gram staining, Tzanck test, darkfield microscopy, slit skin smear, Wood's lamp examination, patch testing, and skin biopsy. KOH examination involves scraping skin and examining under a microscope after applying KOH to identify fungal elements. Gram staining can identify bacteria from infected lesions. The Tzanck test examines blister fluid for signs of herpes or pemphigus. Darkfield microscopy examines exudate for syphilis spirochetes.

2. Skin scraping for KOH
examination
 Involves microscopic examination of stratum
corneum to visualize fungal elements.
 KOH solution causes separation and
destruction of the stratum corneum cells.
 This allows easy identification of exogenous
materials such as hyphae and spores which
are unaffected by the KOH solution.

3. Indications –
 Dermatophytosis of the skin, hair and nails
 Candidiasis
 Tinea versicolor

4. Procedure-
 Swab the site with spirit
 Scrap the lesion at active border with a 15 no.
blade or take hair/nail clipping
 Add 1-2 drops of KOH and put cover slip
 Wait for 15-20 min. for the keratin to digest
(overnight for nail clipping).

5. Dermatophytes- multiple, refractile,
branched, septate hyphae

6. Candidiasis- budding ovoid yeast
cells and pseudohyphae

7. Tinea versicolor- hyphae with
clusters of spores , often called
“spaghetti and meatballs”

8. Scraping for scabies
 After applying a drop of mineral oil, the
burrow is scraped with a 15 no. scalpel blade.
 Scraping transferred to glass slide and seen
under microscope.
 Reveals mite, eggs or fecal pellets.

10. Gram’s staining of exudates
 Used to identify the organism in infected
lesions.
Procedure-
 A thin layer of specimen is spread on a glass
slide, dried, and heat fixed to the glass.
 The slide is flooded with 2% crystal violet and
allowed to stain for 30 seconds to two
minutes and then gently rinsed off with
water.

11.  Incubated in Gram’s iodine for > 30 seconds
(iodine fixes the crystal violet to
peptidoglycans of the Gram-positive cell
wall).
 After rinsing off the Gram’s iodine with water,
the slide is briefly decolorized with acetone.
 Then counterstained with dilute carbol
fuschin for a few seconds, rinsed and air-
dried.

12. A Gram stain of mixed
Staphylococcus aureus
(Gram positive cocci) and
Escherichia coli (Gram
negative bacilli

13. Tzanck test
 Useful in the diagnosis of certain blistering
disorders.
Procedure-
 After deroofing the blister, floor is scraped
and material smeared on a slide.
 Stained with Wright's or Giemsa's stain.

14.  Findings-
 Multinucleated giant cells - diagnostic of
herpes virus or varicella.
 Acantholytic cells- Pemphigus vulgaris
 (Rounded cell with round vesicular nucleus,
perinuclear halo, peripheral condensation of
cytoplasm and lacks desmosomal
connections)

15. Multinucleate
giant cells

16. Acantholytic cell

17. Dark ground microscopy
 The dark ground microscope creates a
contrast between the object and the
surrounding field, such that, the background
is dark and the object is bright.
 Most specific and sensitive technique to
diagnose syphilis when an active chancre or
condyloma lata is present

18. Procedure-
 Lesion is cleaned with saline.
 Its held firmly and pressed b/t thumb and
index finger and the serum exudating is
collected on a cover slip and put on a glass
slide.
 Pressed b/t the folds of a blotting paper to
make a thin, even film.
 Examined under dark ground microscope for
motile treponemes.

20. Slit skin smear examination
 Most important laboratorial test to detect
lepra bacilli in suspected Hansen’s patches
and to classify the d/s.
Procedure-
 Lesion is cleaned with spirit.
 After pinching the skin b/t thumb and index
finger, a 5mm long and 2mm deep cut is
made.

21.  Base is scraped and the material is smeared
on a glass slide.
 After drying and heat fixing the smear, Ziehl-
Neelsen staining is done.

22. Wood’s lamp examination
 Wood’s lamp is a mercury vapor ultraviolet
lamp with a filter which is opaque to all
wavelengths except those b/t 320 to 400
nanometers.
 Mainly emits UV rays of 360 nanometers.

23.  Tinea capitis - yellow green fluorescence
(when the infection is caused by
Microsporum and Trichophyton schoenleini)
 T. versicolor- golden yellow
 Erythrasma- coral red
 Pseudomonas inf.- greenish white
 Vitiligo- milky white
 Albinism- bluish white
 Porphyria- pink / orange (urine)

24. T.capitis
Erythrasma
Vitiligo

25. Vitiligo

26. Patch testing
 Used to identify causes of allergic contact
dermatitis.
Procedure-
 Various patch test allergens (contained within
small metal chambers) are held against the
skin using a paper tape.
 Upper back/ arm

27.  Remains on the skin for 48 hours during
which the person cannot get the tape wet.
 Reading is taken half an hour after removal of
patch.
 2nd
reading at 72 hours.

28.  Allergens used in patch testing include-
metals (e.g. nickel), rubber, leather, hair dyes,
formaldehyde, neomycin,fragrance,
preservative etc..
 Erythema, infilteration, papules and vesicles
indicate positive reaction.

30. Skin biopsy
Process by which a part or whole of the
suspected diseased tissue is obtained for
microscopy and other investigation.
INDICATIONS
 Confirm clinical diagnosis
 Gauge prognosis
 For special investigations
 As a therapeutic modality

31. CONTRAINDICATION
 Bleeding diasthesis
 Active infection at the site
 Keloidal tendency
TYPES
 Shave for exophytic growths
 Punch for endophytic growths
 Excisional for suspected malignancy and as
therapeuticapproach.
 Incisional for deeper lesions

32. Biopsy procedure :
 Select proper site .
 Intradermal or ring anaethesia
 Sample is kept in formalin
 Specimen must be labeled to avoid mixing up
of the slide later..
 Topical antibiotic for one week should be
prescribed .

34. Others
Immunofluorescence –
 Direct
 Indirect
 Used for diagnosis of autoimmune diseases
like- vesicobullous d/s (PV, BP, DH..), SLE,
Lichen planus, vasculities.

35. Serological tests-
 For collagen vascular diseases e.g. SLE,
scleroderma & viral infections and STDs.
Other tests to exclude systemic affection-
 Laboratory tests: CBC, ESR, Liver & kidney
tests, Blood sugar ,urine & stool analysis
 Imaging procedures: U/S, X-Ray.

36. THANK YOU….