Cell and Cell Reaction Flashcards
Best possible resolution for human eye?
200 µm
Diameter of RBC?
7 µm
Fixation in Light Microscopy
10% neutral buffered formalin(formaldehyde stabilized with methanol)
Thickness of cell plasma membrane
0.007 µm thick
Most common stain in light microscopy
Hematoxylin(blue/purple, basophilic) and Eosin(red/pink, acidophilic)
Biopsies
Tissue frozen and frozen sections cut and stained (often with fluorescent tagged antibody)
Transmission Electron Microscopy
An electron beam is transmitted through a thin specimen in a manner similar to the way in which visible light is transmitted through a tissue section for LM
Major difference between TEM and LM
Electron microscopy uses magnetic lenses to focus electrons and LM uses glass lenses to focus photons
Most common fixative in electron microscopy
Glutaraldehyde
How does glutaraldehyde work as a fixative?
forms methylene bridges between polypeptides at reactive side groups
Following glutaraldehyde fixation, tissue must be post-fixed in…
osmium tetroxide to preserve the membrane and lipid components
Dehydration in specimen preparation in TEM
alcohol and acetone and infiltration with epoxy resin
To provide contrast in the microscope of TEM
stained with uranium and lead salts
Scanning Electron Microscope uses
a beam of electron (1° beam) to scan the specimen surface
Main byproducts in SEM as probe scans surface of specimen
secondary electrons, backscatter electrons, x-rays, and photons
Secondary electrons are
low energy electrons emitted from the surface of the specimen and contain surface detail information
SEM and TEM have the same fixation?
True
In SEM, after fixation the specimen is
dehydrated in ETOH and critical point dried
CPD
critical point dried
After drying in SEM, the specimen is
glued onto specimen stub and given a conductive coating (gold,gold-palladium)
Goal of immunocytochemistry
visualize some component(antigen) in a tissue section by means of an antibody usually linked to a fluorescent or colored molecule
Procedure for immunocytochemistry
1)Section tissue 2)Block non-specific binging w/ a protein solution 3)Incubate w/primary antibody and rinse 4)Incubate w/ secondary antibody which contains visualization label
Acid dyes have a __ charge
-
Basic dye has a __ charge
+
Hematoxylin
Blue stain-chromatin (DNA), rough ER, basophilic
Eosin
Pink stain- cytoplasm, collagen/elastin fibers, acidophilic
Standard stain used for tissues in anatomy and physiology
H and E
Periodic acid Shiff stain
Stains glycogen and various carbohydrate containing molecules (magenta)
Basement membrane of Bowman’s capsule in the kidneys stains with
PAS (stains Basement membrane magenta)
Basic structural and functional unit
Cell
Cells are organized into tissue and tissues are organized into
organs
4 primary tissues
Epithelium, Connective Tissue, Muscle, and Nervous tissue
Cytoplasm
Organelles and inclusions suspended in cytoplasmic matrix(cytosol)
Organelles
Cell systems responsible for synthetic, metabolic, energy requiring and energy generating functions of the cell
Thickness of cell plasma membrane
7-10 nm
Plasma membrane functions as a
semipermeable membrane between the cytoplasm and outside of the cell
Membrane is composed of
phospholipids, glycolipids, and cholesterol
Phospholipids possess a polar(hydrophilic) head and two non polar(hydrophobic) fatty acyl tails-so they are called…
amphipathic
Glycolipids are
lipids with attached carbohydrates
Function of Glycolipids
function in cell-cell and cell-extracellular matrix interactions
How are glycolipids distributed
Glycolipids are distributed asymmetrically in outer membrane leaflet
Gangliosides
complex glycolipids with sialic acid(9 C monosaccharide) containing oligosaccharides
GM1 ganglioside
cell surface receptor for cholera toxin
Glycolipids are present only in
outer leaflet and have polar carbohydrate residues that form part of glycocalyx
Glycocalyx
“sugar coat” commonly associated with the extracytoplasmic aspect of the outer leaflet of the plasma membrane
Glycocalyx composition
Polar oligosaccharides side chains covalently linked to protein and some lipid constituents of the plasma membrane
Glycocalyx contain
Proteoglycans, which consist of membrane integral proteins which are bound glycosaminoglycans
Functions of glycocalyx
1)cellular attachment to extracellular matrix components 2)binding of antigens and enzymes to cell surface 3)facilitating cell-cell recognition + interaction
Cholesterol
assists in maintaining membrane structural integrity by decreasing the mobility of the first few CH2 groups on phospholipids.
Cholesterol constitutes
2% of the plasma membrane lipids
Two types of membrane proteins
Integral proteins and Peripheral proteins
Important membrane property
Fluidity
Membrane fluidity increases with
a rise in temperature and by greater unsaturation of the fatty acids in the membrane
Membrane fluidity is decreased by an
increase in cholesterol content
Glucose will move across a RBC membrane down a conc. gradient via
a glucose transporter protein (GLUT1)
GLUT1 is a
multipass transmembrane protein
Mechanism for maintaining ion balance in the cell against an electrochemical gradient
NA+ -K+ PUMP
Sodium and chloride are
10-20x higher outside than inside the cell
Potassium is
20-40x higher inside the cell than outside the cell
NA+-K+ pump mechanism
3NA+ OUT…. 2K+ IN (a single ATP is hydrolyzed to complete two steps)
What inhibits ion transport in the NA+-K+ pump
Oubain- has the same binding site of K+
Sodium-potassium pump acts to
maintain constant cell volume by reducing the ion conc. intracellularly and by increasing it extracellularly- decreasing flow of water into cell
Sodium potassium pump plays a minor role in
the maintenance of a potential difference across the plasma membrane
Movement of sodium down the e-chem gradient can be
coupled to the movement of other molecules against its concentration gradient
Glucose movement across intestinal epithelium works as
cotransported molecule in the same direction as Na+
One reason why tumors are often resistant to a broad range of anticancer drugs
tumor cell over expression of ATPase
Cystic fibrosis transmembrane regulator (CFTR) has a structure closely related to
MDR-ATPase
Cystic fibrosis is due to
defective CFTR in epithelial cell of lung and other tissue
What type of chloride ion channel is CFTR
ATP and cAMP sensitive chloride ion channel
In CF patients, the CFTR
becomes insensitive to cAMP and Cl- flux across the cell membrane
Ion channel proteins
Highly folded transmembrane proteins that form small aqueous pores across membranes - allows specific small, water soluble molecules and ions to pass down gradient
Ion channels influence the rate of flow but not
the direction
Membrane receptors are primarily
glycoproteins located on cell surface that bind specific signaling molecules
Membrane receptors act as
transducers
Membrane receptors control membrane permeability by
regulating the conformation of ion channel proteins
Ion channel linked receptors
binds a signaling molecule that temporarily opens or closes the gate, permitting or inhibiting the movement of ions across the membrane
Catalytic receptors are
single-pass transmembrane proteins. (extracytoplasmic part functions as a receptor and the cytoplasmic part functions as protein kinase)
Catalytic receptors may lack
extracytoplasmic part and therefore be continuously activated (oncogenes)
Transmembrane trimeric G proteins are composed of what 3 polypeptide subunits
alpha, beta ,gamma
