Contemporary Lab Skills - Tissue Culture and Centrifugation

Cells in a determined plastic mobile static( determined, plastic, mobile, static ) state remain the same after differentiating. Most Stem( Most, Stem ) cells are in this state.
Cells in a plastic determined mobile static( plastic, determined, mobile, static ) state are changing, usually before they differentiate. Stem Most( Stem, Most ) cells are in this state.

In order to sanitize using an autoclave, the object must be exposed to 121 115 21 15( 121, 115, 21, 15 ) degrees C at 15 21 115 121( 15, 21, 115, 121 ) psi for 15 10 20 25 30( 15, 10, 20, 25, 30 ) minutes.

Disposal of Biohazardous waste requires some preparation before disposal. Incineration is performed off-site on-site( off-site, on-site ) waste conditioning, while autoclaving is performed on-site off-site( on-site, off-site ).

Substantial Minimal( Substantial, Minimal ) amounts of reduced Nitrogen are required in tissue cultures. Organic nitrogen sources that function better at low high( low, high ) concentrations include glutamine, glutamic acid, urea, and alanine.

Higher Lower( Higher, Lower ) concentrations of auxins and lower higher( lower, higher ) concentrations of cytokinins yield roots.
Lower Higher( Lower, Higher ) concentrations of auxins and higher lower( higher, lower ) concentrations of cytokinins yield shoots.
Other combinations yields callus nothing whole plant growth( callus, nothing, whole plant growth ).

This is a fixed-angle swinging bucket vertical continuous flow( fixed-angle, swinging bucket, vertical, continuous flow ) rotor.

(Type of Separation - Efficiency)
Pelleting - Excellent Poor Good( Excellent, Poor, Good )
Rate zonal - Poor Excellent Good( Poor, Excellent, Good )
Isopycnic - Good Excellent Poor( Good, Excellent, Poor )

This is a swinging bucket fixed-angle vertical continuous flow( swinging bucket, fixed-angle, vertical, continuous flow ) rotor.

(Type of separation - Efficiency)
Pelleting - Inefficient Good Adequate( Inefficient, Good, Adequate )
Rate zonal - Good Inefficient Adequate( Good, Inefficient, Adequate )
Isopycnic - Adequate Inefficient Good( Adequate, Inefficient, Good )

This is a vertical fixed-angle swinging bucket continuous flow( vertical, fixed-angle, swinging bucket, continuous flow ) rotor.

(Type of Separation - Efficiency)
Pelleting - Poor Good Excellent( Poor, Good, Excellent )
Rate zonal - Good Excellent Poor( Good, Excellent, Poor )
Isopycnic - Excellent Poor Good( Excellent, Poor, Good )

This is a continuous flow fixed-angle vertical swinging bucket( continuous flow, fixed-angle, vertical, swinging bucket ) rotors.

(Type of separation - Efficiency)
Pelleting - Poor Excellent Adequate( Poor, Excellent, Adequate )
Rate zonal - Excellent Poor Adequate( Excellent, Poor, Adequate )
Isopycnic - Adequate Poor Excellent( Adequate, Poor, Excellent )

Centrifugal rotors are rated for maximum operating speed size average operating speed user satisfaction( maximum operating speed, size, average operating speed, user satisfaction ). The rating is established by the manufacturer a scientific commitee( the manufacturer, a scientific commitee ) based on construction materials height maximum number of samples( construction materials, height, maximum number of samples ) and design radius balance( design, radius, balance ).

De-rating is the reduction in maximum operating speed time frequency( speed, time, frequency ) after a period of use. The de-rating schedule is established by the manufacturer the user a scientific committee( the manufacturer, the user, a scientific committee ). It prolongs the useful life of the rotor, albeit at a lower maximum operating speed frequency time( speed, frequency, time ).

The picture depicts pelleting rate zonal centrifugation isopycnic centrifugation( pelleting, rate zonal centrifugation, isopycnic centrifugation ), which happens when using a fixed angle vertical swinging bucket continuous flow( fixed angle, vertical, swinging bucket, continuous flow ) rotor to centrifuge until the particles sediment to the bottom of the tube.

Differential pelleting Rate zonal centrifugation Isopycnic centrifugation pelleting( Differential pelleting, Rate zonal centrifugation, Isopycnic centrifugation, pelleting ) happens after repeating centrifugation at higher RCFs. The pellets sediment to the bottom based on size and mass
(Nuclei Chloroplast Mitochondria( Nuclei, Chloroplast, Mitochondria ) > Chloroplast Nuclei Mitochondria( Chloroplast, Nuclei, Mitochondria ) > Mitochondria Nuclei Chloroplast( Mitochondria, Nuclei, Chloroplast ))

This is rate zonal centrifugation pelleting differential pelleting isopycnic centrifugation( rate zonal centrifugation, pelleting, differential pelleting, isopycnic centrifugation ), which separates particles based on mass and shape. The sample is stabilized by a sucrose glucose( sucrose, glucose ) gradient so that sedimentations happen in layers (sedimentation constants proportional to molecular weight size density( molecular weight, size, density )).

A swinging bucket vertical fixed angle continuous flow( swinging bucket, vertical, fixed angle, continuous flow ) rotor is used for analytical purposes and small-scale preps, while a vertical fixed angle swinging bucket continuous flow( vertical, fixed angle, swinging bucket, continuous flow ) rotor is used for larger particles.

This is isopycnic centrifugation pelleting rate zonal centrifugation differential pelleting( isopycnic centrifugation, pelleting, rate zonal centrifugation, differential pelleting ), or equilibrium density gradient centrifugation. In this kind of centrifugation, migration stops when particle density heat energy( density, heat energy ) equals medium density heat energy( density, heat energy ).

For this kind of centrifugation, a vertical swinging bucket fixed angle continuous flow( vertical, swinging bucket, fixed angle, continuous flow ) rotor is fast and produces high resolution. A fixed angle vertical swinging bucket( fixed angle, vertical, swinging bucket ) rotor produces a high resolution, and a swinging bucket vertical fixed angle( swinging bucket, vertical, fixed angle ) rotor has a high capacity.

Label as it applies to the pictured centrifugation (isopycnic fixed angle continuous flow swinging bucket( isopycnic, fixed angle, continuous flow, swinging bucket )):

Typical gradients:
CsCl Percoll Ca2SO4( CsCl, Percoll, Ca2SO4 ) (most common)
Percoll Ca2SO4 CsCl( Percoll, Ca2SO4, CsCl ) (common)
Ca2SO4 CsCl Percoll( Ca2SO4, CsCl, Percoll ) (occasionally)

Typical starting density = 1.55 1.15 5.11 5.51( 1.55, 1.15, 5.11, 5.51 ) CsCl/mL

Density or buoyant density (p) in CsCl
Protens = 1.3 1.16 1.8( 1.3, 1.16, 1.8 )
dsDNA = 1.6-1.7 1.3-1.6 1.75-1.8( 1.6-1.7, 1.3-1.6, 1.75-1.8 )
RNA = 1.75-1.8 1.3-1.6 1.6-1.7( 1.75-1.8, 1.3-1.6, 1.6-1.7 )

p is used to separate and characterize DNA RNA Chloroplasts( DNA, RNA, Chloroplasts )
p = 1.660 + 0.098*(G + C)
where (G + C) = mole density( mole, density ) fraction

The picture depicts a gradient maker Hamilton machine gradient chamber( gradient maker, Hamilton machine, gradient chamber ): a two three four( two, three, four )-chambered apparatus that controls the mixing of two solutions to form a gradient.

One way of concentrating samples is through precipitation spin collumns( precipitation, spin collumns ), where one centrifuges to pellet and collects the precipitate. Then one resuspends the pellet in the buffer to the desired concentration.

Nucleic Acids Proteins( Nucleic Acids, Proteins )
- Ethanol
- Isopropanol
Proteins Nucleic Acids( Proteins, Nucleic Acids )
- Acetone
- Ammonium sulfate
- Isopropanol
- Trichloroacetic acid

One way of concentrating samples is spin columns precipitation( spin columns, precipitation ), where one centrifuges a tube with 2 chambers with a molecular weight density size selection( molecular weight, density, size selection ) cut off membrane separating the compartments. Before centrifugation, the upper chamber contains a dilute concentrated( dilute, concentrated ) sample. The lower chamber collects the filtrate concentrated sample( filtrate, concentrated sample ) during centrifugation, while the lip surrounding the membrane collects the concentrated sample filtrate( concentrated sample, filtrate ) in the upper chamber.