Staining Techniques Flashcards
staining of these is very difficult because they hardly absorb, reflect ,refract or diffract much light
microorganisms
uses one dye and can be positive or negative staining
simple staining
uses basic dyes
positive simple staining
-uses acidic dyes
negative staining
– uses two or more dyes that will distinguish bacteria from each other because their cell walls have different reactions on dyes
differential staining
technique to add stain to specimens to make it more visible under the microscope
staining technique
– are of little use unless the specimens for viewing are prepared properly
microscope
important tool that aids in the observation of microorganism
microscopy
- is a culture-free method to identify and compare bacterial diversity from complex microbiomes or environments that are difficult to study
16s rRNA sequencing
– have the same refractive index as water
microorganism
When observed under a microscope, they are transparent or nearly invisible to the naked eye
microorganism
Must be fixed and stained to increase visibility, accentuate specific morphological features, and preserve for future use
microorganism
substance that adheres to a cell, giving the cell color
stain
Presence of color gives the cells significant contrast so are much more visible
stain
They are used to differentiate different types of organisms or to view specific parts of organisms
stain
spherical bacteria
cocci
rod shaped bacteria
bacilli
helical, twisted, spirochete shaped bacteria
spirilla/spirillus
comma shaped bacteria
vibrio
view different bacteria shapes
+1
- Bacterial arrangement (4)
clusters
chains
pairs (diploids)
no special arrangement
view different arrangement of cocci
+1
arranged in pairs or chains
strep
clusters, pairs and occasionally in short chains.
staphylococci
involves directly staining the bacterial cell with a positively charged dye in order to see bacterial detail, in contrast to negative staining where the bacteria remain unstained against a dark backround
simple staining
positive dye is used to adhere to a negatively charged plasma membrane (ex. Crystal violet, methylene blue)
positive staining
uses basic dyes to color the specimen against a bright background
positive stianing
the type of chromophore used in this technique is a positively charged ion instead of a negative one
positive staining
– utilizing negative dye (nigrosin)
negative staining
performed for the observation of intact microbial structures without disturbing its cellular morphology
negative staining
the bacteria cells are not stained, rather the glass background containing cells.
negative staining
procedure where more than one dye is used to differentiate between different types of microorganisms on a slide
differential staining
Helps differentiate between cell types and cell structures
differential staining
differential staining procedure used to categorize bacteria as Gram-positive or Gram-negative based on the chemical and physical properties of their cell walls.
gram staining
- is a differential stain used to identify acid-fast organisms such as members of the genus Mycobacterium. (+ is pinkish to reddish)
acid fast staining
introduced differential staining called gram staining.
Hans Christian Gram
– has an outer lipid membrane that does not adhere with crystal violet, most likely have a flagellar structure
gram negative bacteria
– have a thicker peptidoglycan cell wall, most likely to be toxin producing
gram positive bacteria
– bacteria stain different colors depending on their cell wall construction
differential stain
– thick peptidoglycan layer , lacks an outer lipid membrane
gram positiv bacteria
– have an outer lipid membrane and a thin peptidoglycan cell wall
gram negative bacteria
groups of bacteria that lack the protective cell wall peptidoglycan structure
mycoplasma
do not have peptidoglycan layer
archaebacteria
- The difference in cell wall construction effects
Which antibiotic will be more effective
susceptibility to disinfectants
toxins produced
toll-like receptors recognition
flagellar structure
have linear glycan strands cross-linked by short peptides wherein the glycan strands
structure of peptidoglycan
peptidoglycan are cross linked by what
short peptides
glycan strands in peptidoglycan is made out of
n-acetylglusoamine
n-acetylmuramic acid
- gram staining procedures step 1
make a smear and heat fix
- gram staining procedures step 2
primary stain (crystal violet) for 1 minute then rinse
primary stain
crystal violet
- gram staining procedures step 3
mordant (gram’s iodine) for 1 minute then rinse
what is the mordant for gram staining
gram’s iodine
binds to the crystal violet making a large complex that adheres to the cell membrane
gram’s iodine (mordant)
- gram staining procedures step 4
add decolorant
decolorant is what
95 ethanol or acetone
removes the crystal violet from Gram-negative cells since crystal violet did not adhere strongly to the thin peptidoglycan layer and the cells become transparent
decolorant
- gram staining procedures step 5
rinse with water immediately
- gram staining procedures step 6
counterstain (safranin) for 1 minute
as a counterstain to stain transparent cells (Gram-negative cells) pink
safranin
- gram staining procedures step 7
blot dry and view with microscope
– helps in visualization of a bacterial structure. This staining is crucial as some bacterial structures play the role of endotoxin and antigenic
structural staining
spores that are formed within the cells, mainly in bacteria. These spores are formed within bacteria vegetative cells or mother cells known as sporangia.
endospore
nutrients that are supplied to microorganisms are in the form of
culture medium
aqueous solution of nutrients that contains energy source, macroelements, trace elements, and growth factors
medum
formulated so that the concentration and chemical natuer is known
chemically defined medium
intended to remove forms of life
sterilization
