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UND 362 instrumentat
UND 362 instrumentation
| Question | Answer |
|---|---|
| Name the 9 parts of a microscope | bi/monocular eyepiece, objectives (4x etc.), stage, light source, rough focus, fine focus diaphragms, stage dial controls, dimmer switch (light on/off/intensity) |
| name the 9 standard steps of histolgy | accessioning, grossing, processing, embedding, microtomy & drying, stainer, cover slippihng, special staining, review of slides |
| List 4 common things that are done during accessioning | case #'s given, verification of spec types, specimen triage (split for flow/micro), labelling of cassettes and slides |
| name three types of specimens during accessioning | fixed, unfixed (FS), cytology 1. smears (bronch wash/brush 2. cell block (concentrate spec via centrifuge 3. FNA |
| list 4 common things done during "Grossing" | spec prep - fixing, decal, electron microscope prep, quantitative analysis |
| what are the steps in processing (closed system) | fixation (formalin), dehydration (increasing % conc of alcohol, clearing with xylene, infiltration with molten paraffin |
| What takes place during embedding | specs put in molten wax (in specific orientation), additional wax is added and then cooled on cold plate |
| what are 4 common principles of a microtome | most are rotary (moved in 4-6 microns, hold block firmly, sections tissue thinly, created ribbons |
| What are 4 ways slides can be dried | air dry (helps avoid artifacts via lower temp), dryer, incubator, microwave |
| H/E staining positive and drawbacks to manual and automatic processing | manual- inexpensive, reusable containers, but need QC Auto-high volume, need QC but is consistent |
| Coverslipping pos/negs to manual and auto | manual-labor intensive, inexpensive, can lead to chem exposure auto-need daily QC, high throughput |
| What can special stains do/types | ID specific tissue components: carbohydrates, amyloid, nerve, pigments, granules, minerals |
| IHC (Immunohistochemistry) used two types of modes what are they | 1. antibody to antigen attraction for staining (markers for the inclusion or exclusion of a diagnosis 2. In Situ hybridization- use gene probe to locate specific dna/rna and color with chromagen |
| list 6 types of microscopes | light, polarizing, phase contrast, darkfield, flourescence, electrom (TEM -transmission, SEM-scanning) |
| list the 4 typical objectives of a light microscope | 4x (scanning), 10x(intermediate), 40x(hidry), 100x (oil immersion) (the oculars are 10x) |
| total magnification is determined by | oculars (10x) x the objective used = |
| Koehler illumination is also available on the light microscope but needs? | additional optics |
| what is a polarizing microscope used for? | ID of crystals (talc, sylica, urates) |
| Birefringence and double refraction is? | light ray will be split into 2 that emerge from the crystal at different points |
| how is polarizing achieved | polarizer between spec and light source, and another polarizer filter between spec and eye |
| what is polarized light commonly used to see? | tissue stained with congo red (seen as apple green birefringence) |
| What is a phase contrast microscop commonly used for? | for unstained specs and living specs such as protozoa. |
| What would a darkfield microscope be used for | unstained microorganisms (spirochetes) |
| how is a darkfield micrscope used | direct light is excluded and oblique light (light from an angle) is used |
| Flourescence microscopes are used for what | visualization of flourescent antibodies |
| give 2 primary examples of spec/stain combo that is used with a fourescent microscope | acid-fast bacillus w/ auramine rhodamine stain amyloid w/ thioflavin T staining method |
| How is flourescence achieved and what does the microscope require? | light wavelength is absorbed and reemitted as a longer one (typically uv and emitted as visible light) REQUIRES mercury or halogen lamp |
| name 2 common flourescence dyes | Fluorescein isothiocyanate, rhodamine |
| how is microscope set up for flourescence | exciter filter between light and spec transmits desired light wave(not visible light), filter put in eyepiece to absorb UV and only allow visible light |
| Give 5 specifics for TEM (transmission electron micro) | only in large institutions, use beams of electrons in vacuum, 1000x to 500,000x, requires special fixative, use heavy metal stains |
| What and how is the SEM (scanning elecron micro) used | Surface or 3d imaging, back scattered electrons are picked up for image |
| describe and give 1 pos. and a 2 disadv. for open processors | beakers with reagent(s) POS custom design, NEG evaporation and safety issues |
| Explain how a closed processor functions | tissue in resivour and solutions are pumped in and out |
| give 6 adv to closed processor | tissue can't dry out, QC done by instrument, many programs/schedules, heat/vacuum/agitation in one place, alarm system, safety (minimal or no vapors) |
| give 2 disadv to closed processors and 4 QC properties | can't/may not be able to use all reagents, heat and vacuum must be used carefully. QC frequent reagent rotation, fluid above cassettes, paraffin 2-4○ above melt pt., document qc |
| describe microwave tissue processors and give 3 adv | closed system, only large instiutions, expensive ADV- shortens processing time, avoids xlyene use, used only 4 reagents after preprocessing solution |
| what is the melting point of paraffin in the embedding station | 2-4○ above melting point |
| name the 5 types of microtomes | rotary, cryostat, sliding, ultra, clinical freezing |
| explain rotary microtome | most common, hand wheel moves block forward typicall 4-5 microns, routine cleaning needed |
| what is the typical configuration of the cryostat and its normal temp | chill chamber with rotary microtome, 10-25○ below 0○C |
| what is the typical knife point for the cryostat | 30○ (w/ wedge shaped knife) |
| what is a artifact to avoid with cryostats | ice crystals can form if not frozen quickly enough (can use liquid nitrogen to snap freeze which will avoid crystals and therefore holes in the sample) |
| what is the QC for the cryostat | universal precautions, keep clean, decontaminate |
| explain how/what a clinical freezing microtome is | portable cryostat of sorts (uses constant CO2 to freeze)- block is moved up and the knife moves horizontally above it |
| what are 2 drawbacks to the clinical freezing microtome? | poor for friable tissue, airborne debris may result from the C02 |
| how is the sliding microtome used and for what | used for research (block and knife are horizontal) - used on celloidin and large paraffin blocks |
| what type of knives are used in the ultra microtome? | diamond or glass knives |
| what it he ultra microtome used for and how | cuts .5 micrometer sections for light microscope and 90nm (ultra thin) for eletron microscope (requires training) |
| what are the four types of microtome knives | steel, disposable, glass, diamond |
| what are the three shapes of microtome knives | wedge, planoconcave and biconcave |
| What are the wedge/planoconconcave/biconcave knife shapes used for | wedge - paraffin, carbowax, frozen, celloidin plano- celloidin biconcave - rarely used |
| what is the clearance angle on a microtome? wedge angle? cutting facet angle? | 3-8, 15-18, 27-32 |
| give three facts about water baths | allow paraffin to decompress, high heat causes parched earth, kept at 5-10○ below melting point of paraffin (stretch ribbon gently) |
| what are 6 types of slide adhesives | gelatin, albumin, glue, poly L lysine, charged slides, silane |
| how is a slide dryer typically used | dries paraffin off sections of slides (typically set just above paraffin melting point around 60○) |
| name the 4 types of stainers | manual, linear, revolving, robotic (L/R/R are expensive) |
| name two types of special stainers | Immunohistochemistry (IHC and In Situ), auto "routine" special stainer |
| describe the method for manual staining | use coplin jars in sequential order (drying, deparaffin, rehydration, dye uptake, counterstain, clearing, coverslip) |
| describe linear stainers | transfers slices from one container to the next, can be continually loaded, staining times can ONLY be varied by the # of containers (of each chem), dries the slides |
| what is the main difference between linear and revolving stainers | they are the same but revolving can have its times varied |
| what are 3 descriptions of a robotic stainer | computerized, some do the drying, no particular order for slides since the stainer knows where they need to go |
| what are 2 properties and 2 adv. of auto coverslippers | must be cleaned and QC/glass and plastic covers ADV. hight output and reduces xylene exposure |
| regarding ph if acid is added what will occur what if a base is added | acid = hydroniom ion conc. increase = lower ph base = hydroxy ion conc. increase = higher ph |
| how do you calibrate a solution | use a standard (known spec) that is close to the solution to be calibrated |
| what temp are incubators typically set at and what are they used for | 37○, enzyme stains (not recommended for slide drying) |
| what are the typical temps for lab fridges and freezers | fridge (2-10○C), Freezer (-20○C) |
| what can fridge and freezers be used for? | storing aliquots (acetone and ether must be in explosion proof fridge/freezer) |
| what is a hydrometer used for | measures specific gravity -or- % alcohol |
| define koehler illumination | extremely even illumination of a sample (needs additional optics) |
| what is auto flourescence | naturally occuring fourescence (ie collagen fibers) |
| define achromatic, apochromatic, parfocal | achromatic (corrected for 2 colors) apochromatic (corrected for 3 colors and other lens aberrations) parfocal- all objectives have focal point in same plane and magnification |
Created by:
mustangvxd
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