Falcon tube

Tests were carried out at 25°C and at initial pH 6.9. Cultures in the liquid medium were incubated in 50 mL Falcon tubes, continuously shaked at 220 rpm. Each culture contained a fresh Pseudomonas sp. 0X1 colony in 10 mL of medium. The airlift with 10 g of pumice was sterilized at 121°C for 30 min and then housed in a sterile room. One-day culture was transferred to the reactor and, after a batch phase, liquid medium with phenol as the only carbon source was continuously fed. The reactor volume V was fixed at 0.13 L. Aerobic conditions were established sparging technical air. Under these conditions microorganism started to grow immobilized on the solid s support. When immobilized biomass approached steady state, cyclic operation of the airlift was started by alternating aerobic/anaerobic conditions. 

The reaction was quenched by transferring it into a 50 mL Falcon tube containing 33 mL of ice-cold acidic methanol and left immersed in ice for 20 min. The proteinaceous pellet was precipitated by centrifugation at 8000 rpm for 15 min at 4 °C and the supernatant collected in a glass bottle. The pellet was re-extracted twice by washing it in MilliQ water (5 mL) and reprecipitated with ice-cold 100% methanol (35 mL) over 20 min in ice and then centrifuged as above. The combined supernatants were evaporated at 35 °C and the residue was reconstituted in 20 mL of mobile phase (4—8 % MeCN in 5 him ammonium acetate, pH 5.0). 

L-shaped spreaders, plastic, sterile (e.g. VWR1612-1560, VWR International) two polypropylene tube, 50 mL, screw-capped, presterilized (e.g. Falcon tubes, Becton Dickinson Labware, Franklin Lakes, NJ, USA)... 

A spore suspension was prepared by resuspending the spores from the agar plate in 15 mL physiological aqueous solution (8.5 g NaCl, 1 g peptone, and 1 g Tween 80 in 1 L distilled water, sterilized at 121 °C for 20 min) with a Drigalski spatula. The spore suspension was diluted to a concentration of approximately 2.5 x lO spores/mL and stored in a 50 mL Falcon tube at 4 °C. 

Cool the samples to RT and transfer the content to 15 ml falcon tubes. 

Collect the mesh flow through harboring the cells and transfer it into a fresh sterile falcon tube. 

Take 20ml of ficol (Pharmacia) in a 50ml Falcon tube (5 tubes). 

A single colony is inoculated into 20 ml of GS96 medium with 1 % w/v glucose and antibiotics in a 50 ml falcon tube. The culture is incubated at 37°C and 225 rpm overnight ( 16 hours). 

Transfer the wet gel into a botde (e.g., 50-ml Falcon tube), and add the ligand dissolved in 25 ml of Soln. B. 0.5-1 mmoles or 5-10 mg of protein per milliliter are optimal. Less ligand is also possible. If only few ligand is available, reduce the amount of gel and buffer. 

Mix well the contents of the Ficoll bottle, wipe the rubber cap with 70% ethanol then inject approx 15 mL air with a 20-mL syringe and needle and withdraw 20 mL of the Ficoll solution. Equally divide the Ficoll solution between two new 50-mL Falcon tubes. 

Using a 10-mL pipet and pipetor, carefully aspirate the plasma layer from the Ficoll tubes and discard. Leave approx 5 mL of plasma on top of the mononuclear cell layer to avoid cell loss. Carefully aspirate the mononuclear cell layer and transfer to two new 50-mL Falcon tubes. A total of approx 12 mL of the mononuclear cell volume should be obtained. Move the pipet tip in a circular, continuous manner over the mononuclear cell layer when aspirating avoiding red blood cell contamination. Discard the red blood cell contents in to the waste container containing 10% bleach solution. 

Resuspend cells by flicking tube and transfer into 5 ml of HT-29LP medium with geneticin in a 50 ml polypropylene Falcon tube. 

Pellet cells for 5 min in two 50 mL Falcon tubes (e.g., 2000 rpm in a Beckman J-6M swinging bucket centrifuge with Beckman JS-4.2 rotor). (We use sterile Falcon tubes (BD Biosciences, San Jose, CA, cat. 352070) to minimize the chance of nuclease contamination.) The supernatant is removed. Optionally, one pellet can be resuspended in equal volume of LB (1% Bacto Tryptone, 0.5% yeast extract and 0.5% NaCl in water) and 60% glycerol, divided into multiple fractions and stored at —80 °C for future use. The other pellet is resuspended in 25 mL at 37 °C LB, then add into 2 L LB containing 2 mL of 50 mg/mL Carbenicillin and 2 mL of 10 mg/mL Chloramphenicol. Grow at 37 °C until the ODgoo is close to 1,... 

Ice-cold ethanol 50-ml Falcon tube Tooth picks... 

Note The ethanol must be at least po% and it needs to be cold. Using a plastic pipette makes it easy to dispense. Cut squares of cheesecloth (two layers thick) large enough to hang over the edge of the Falcon tube. This activity can be completed in one 40-minute class period. 

When you re finished, place the cheesecloth over the Falcon tube. 

Mountant for fluorescence microscopy can be obtained from several commercial sources, but we make our own. 2.4 g Moviol (Calbiochem) are added to 6 g analytical grade glycerol and 6 mL H20 in a 50-mL Falcon tube at room temperature. The dissolution of the Molviol is completed by the addition of 12 mL 0.2 M Tris-HCl, pH 8.5, and the solution is incubated at 50°C for 10 min. The solution is clarified if necesssary by centrifugation at 5000g for 15 min. The solution is stored at -20°C in suitable aliquots. 

The resultant cell suspension was then transferred into 15-mL Falcon tubes, washed with IX PBS or RPMI, and the cells were counted. 

Filter into a 15-mL Falcon tube by washing the support with additional TFA (0.5 mL). Concentrate the filtrate to approx. 200 pL and precipitate the PNA oligomer with cold (4°C) diethyl ether. 

Decant supernatant into 50 mL falcon tubes (conical bottom) and spin at 1000,g room temperature, 10 min. 

Trypsinize cells as normal and transfer to a 50-mL falcon tube. 

PAXgene tubes placed directly from room temperature to -80 °C tend to crack on thawing, and can be placed in to 50 ml falcon tubes to prevent loss of sample and contamination of lab surfaces. This cracking is reduced if samples are stored overnight at -30 °C before being placed in a -80 °C freezer. 

To create an ice bath for the sonication of lipids (to prevent heat-induced chemical degradation of lipids), fill a 50-mL Falcon tube three-fourths full of ice and pour in water to fill in the gaps between the pieces of ice. 

Mix 89.6 pi of 5 M sodium hydroxide solution, 140 pi of 200 mM vanadate solution, and 2,570 pi sodium carbonate solution in a 15 ml disposable round-bottom Falcon tube, followed by thorough mixing by vortexer. This is the Color Developer referenced in subsequent steps, and it should be prepared fresh every assay day. The pH value should be between... 

Mix 2,000 pi of 1 M Tris solution, pH 7.5,40 pi of 1 MMgCl2 solution, 16 pi of 50 mM ZnCU solution, 4,603 pi of 13.5 mM pNPP solution, and 3,341 pi water in a 15 ml round-bottom Falcon tube, followed by thorough mixing by vortexer. Transfer 3 ml into another round-bottom Falcon tube and add 150 pi of 200 mM vanadate solution and mix by vortexer. These two tubes contain the assay mix and control mix, respectively. 

Transfer 3 ml into another round-bottom Falcon tube and add 150 [il vanadate solution and mix by vortexer. These two tubes contain the assay mix and control mix, respectively. 

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