Troubleshooting

Question 1

H&E error: Tiny dart granules on top of tissue

Answer 1

Cause/Resolution:

Hematoxylin precipitate - filter

Question 2

H&E error: Dull red/brown nuclear staining

Answer 2

Cause/Resolution:

Overripe hematoxylin or insufficient bluing

Question 3

H&E error: Nuclei with blue ring and colourless center

Answer 3

Cause/Resolution:

Stain time too short

Question 4

H&E error: Uniform weak staining of nuclear membrane

Answer 4

Cause/Resolution:

over differentiation

Question 5

H&E error: Blue, congested nuclei, no proper chromatin detail

Answer 5

Cause/Resolution:

insufficient differentiation

Question 6

H&E error: Overall muddy purple appearance

Answer 6

Cause/Resolution:

acidic formalin

Question 7

H&E error: Pale pink

Answer 7

Cause/Resolution:

slow dehydration - retain in eosin and dehydrate more rapidly

Question 8

H&E error: Increase in eosin staining

Answer 8

Cause/Resolution:

Not rinsing slides adequately following bluing

Question 9

Mucicarmine error: Staining with mucicarmine masked

Answer 9

Cause/Resolution:

overstained with nuclear or counterstain

Question 10

Alcian blue error: Weak staning

Answer 10

Cause/Resolution:

Poor hydration - increase hydration time

Question 11

Alcian blue error: Nuclei stain

Answer 11

Cause/Resolution:

overstaining

Question 12

Congo red error: Decreased staining

Answer 12

Cause/Resolution:

Prolonged fixation in formalin

Question 13

Argyrophilic error: Over impregnation of reticular fibres

Answer 13

Cause/Resolution:

new section

Question 14

Argyrophilic error: decreased reticular staining

Answer 14

Cause/Resolution:

Slides washed for too long between impregnation and reduction steps

Question 15

Von Kossa error: Weak staining

Answer 15

Cause/Resolution:

using acids prior to staining

Question 16

Von kossa error: False positives

Answer 16

Cause/Resolution:

Melanin and argentaffin granules present

Question 17

Toluidine blue error: weak staining

Answer 17

Cause/Resolution:
Alcohol destroys polymer function as dye is small and easily diffuses and decreases metchromasia
Blot slides

Question 18

Toluidine blue error: increased stain color

Answer 18

Cause/Resolution:

decrease stain time

Question 19

Toluidine blue error: darker staining

Answer 19

Cause/Resolution:

occurs with elevated pH, make sure pH is close to 4

Question 20

Weigert Van Gieson error: Dull overall stain

Answer 20

Cause/Resolution:

unsaturated picric acid

Question 21

Massons trichrome error: poor staining with red cytoplasmic stain

Answer 21

Cause/Resolution:
overuse of stain, aging of stain
discard and replace stain

Question 22

Masson trichrome error: Muscle stains purple

Answer 22

Cause/Resolution:

not using Bouins, or it is too old

Question 23

Modified argentaffin error: nonspecific staining

Answer 23

Cause/Resolution:

Solution breakdown due to overheating methenamine silver

Question 24

Prussian blue error: False positives

Answer 24

Cause/Resolution:
preparing reagents with tap water (Must use iron free dH2O)
Metal containers, forceps
Must use reagent or annular grade HCl (may contain iron otherwise)

Question 25

Prussian blue error: diffuse background reaction

Answer 25

Cause/Resolution:

residual iron containing solution not completely removed during glassware cleaning