Structures appear shrunken |
Improper fixation |
Alter strength or type of fixative |
Hypertonic buffer |
Adjust salt concentration |
Excessive holes in membranes |
Inadequate dehydration |
Start dehydration series with lower concentration of ethanol or acetone |
Specimen exposed to air during fluid exchanges |
Avoid unnecessary exposure to air. |
Avoid complete removal of solutions during exchange, do more exchanges |
Inadequate exchange in critical point dryer |
Do more and extended exchanges of fluids in the CPD |
Put under vacuum before completely dried |
Allow specimen to dry overnight before placing under vacuum |
Debris on specimen |
Incomplete washing with buffer |
Additional wash steps prior to first fixations may be necessary |
Use of mild detergents or other reagents may be required |
Capsular material appears sloughed off in background |
Inadequate fixation |
Additives may be required for stabilization, i.e. alcian blue or ruthenium red. |
Adjust pH or buffer appropriately |
Specimen moves while imaging |
Poor contact with mounting substrate |
Add conductive paint around specimen to “seal the cracks” |
Not enough material adhering to substrate |
Concentration of specimen too low |
Allow longer settling time before fixation |
Increase initial concentration |
Poor adherence properties |
Pre-treat mounting substrate with adhesive |
Excessive charging of sample (SeeTable 4)
|
Poor contact of specimen to mount |
Improve contact by more firm placement on conductive adhesive, and coat with conductive paint if appropriate |
Inadequate metal coating |
Increase thickness of metal coating |
Inadequate bulk conductivity |
Increase bulk conductivity through OTOTO |
Fine detail seems obscured And/or surface appears “chunky” |
Excess metal applied during sputter coating |
Apply thinner layer of metal |
Use finer grain or lower atomic number element |