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.

Indications –

Dermatophytosis of the skin, hair and nails Candidiasis Tinea versicolor

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).

Dermatophytes- multiple, refractile, branched, septate hyphae

Candidiasis- budding ovoid yeast cells and pseudohyphae

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

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.

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.

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.

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

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.

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)

Multinucleate giant cells

Acantholytic cell

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

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.

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.

Base is scraped and the material is smeared on a glass slide.

After drying and heat fixing the smear, Ziehl-Neelsen staining is done.

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.

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)

T.capitis

Erythrasma

Vitiligo

Vitiligo

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

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.

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.

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

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

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 .

Others

Immunofluorescence – Direct Indirect

Used for diagnosis of autoimmune diseases like- vesicobullous d/s (PV, BP, DH..), SLE, Lichen planus, vasculities.

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.

THANK YOU….