K. veneficum strain CCMP 2064 that produces Kmtx2 was acquired from the Center for the Culture of Marine Phytoplankton (CCMP). CCMP 2064 was isolated in November 1998 from a fish-kill on the Wilmington River, Georgia and deposited on May 3, 2003. CCMP 2064 is unialgal but not axenic and was cultured phototrophically in 15 psu filtered (0.22 μm) natural seawater combined with f/2 nutrients and vitamins without Si−1 at 100 Einstein m−2s−1 and 20°C ambient temperature. Cell densities were measured using a Coulter Multisizer II (Beckman-Coulter) counter and cells mL−1 determined using Accucomp (Version 2.01) software. Abbotoxin and 59-E-chloro-Abbotoxin were isolated from the type species (PLY 103) for K. veneficum originally isolated in 1950 from Plymouth Sound, UK (50.364 N, 04.182 W). PLY 103 was grown in 500 ml glass flasks with 300 ml of culture in ERD-SCHREIBER medium at 15 degrees and 12:12 light:dark cycle. ambient temperature. Richard Pipe of the MBA initially grew 3 liters of PLY 103 (~19, 700 cells/ml; ~6 × 107 cells). The cultures were filtered on GF/F filters and the filters and frozen filtrate on dry ice were sent to Maryland. Subsequently, he grew 14 liters of PLY 103 grown in Erd-Schreiber Media containing 50 mg NaH13CO4 (sodium bicarbonate) and placed the filtrate on a 55g C18 columns (AnaLogix, Sorbtech Technologies) for shipping to Maryland.
Coulter multisizer 2
Manufactured by Beckman Coulter
Sourced in United States
The Coulter Multisizer II is a laboratory instrument used for particle size and particle count analysis. It utilizes the Coulter Principle to accurately measure the size and concentration of particles suspended in an electrolyte solution.
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3 protocols using coulter multisizer 2
1
Culturing and Isolating Toxins from Karenia veneficum
2
Algae Growth Conditions Protocol
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All species were grown under normal growth conditions as follows. Chlorella variabilis NC64A and Coccomyxa subellipsoidea cultures were grown in modified Bolds basal medium (MBBM; Van Etten et al. 1983 (link)) shaken at 100 RPM, 22°C, and a light intensity of 30 µE. Chlorella sorokiniana UTEX 1230 was obtained from the University of Texas Culture Collection and grown in liquid Bolds Basal Medium (BBM; Nichols and Bold 1965 ), shaken at 115 RPM, 25°C, and a light intensity of 58 µE. Chlamydomonas reinhardtii CC124, obtained from Dr. Donald Weeks, was grown in Tris‐Acetate‐Phosphate medium (Gorman and Levine 1965 (link)), shaken at 100 RPM, 22°C, and a light intensity of 30 µE. Cell abundance was determined using a Coulter Multisizer II instrument (Beckman Coulter, Fullerton, CA, USA).
3
Characterization of Graphene Nanoparticles
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The average particle size
(d50) of graphene was around 1377.5 ±
140.8 nm, which was measured by a particle size analyzer (Coulter
Multisizer II, Beckman Coulter, Fullerton, CA). The surface area of
graphene was measured to be 212.56 m2/g by using a surface
area analyzer (Coulter SA 3100, Beckman Coulter, Fullerton, CA). The
zeta-potential of graphene at pH = 7.0 ± 0.2 was around −19.98
eV by a zeta-potential analyzer (Coulter DELSA 440SX, Beckman Coulter,
Fullerton, CA). The X-ray diffraction pattern of graphene was recorded
by an X-ray powder diffractometer (D8 ADVANCE, Bruker, Germany) (Figure S1 ). The morphology of graphene was examined
by scanning electron microscopy (SEM) (Hitachi S-4800 FEG SEM, Tokyo,
Japan) (Figure S2 ) and transmission electron
microscopy (TEM) (Philips Tecnai G220 S-TWIN, Amsterdam, The Netherlands)
(Figure S3 ).
(d50) of graphene was around 1377.5 ±
140.8 nm, which was measured by a particle size analyzer (Coulter
Multisizer II, Beckman Coulter, Fullerton, CA). The surface area of
graphene was measured to be 212.56 m2/g by using a surface
area analyzer (Coulter SA 3100, Beckman Coulter, Fullerton, CA). The
zeta-potential of graphene at pH = 7.0 ± 0.2 was around −19.98
eV by a zeta-potential analyzer (Coulter DELSA 440SX, Beckman Coulter,
Fullerton, CA). The X-ray diffraction pattern of graphene was recorded
by an X-ray powder diffractometer (D8 ADVANCE, Bruker, Germany) (
by scanning electron microscopy (SEM) (Hitachi S-4800 FEG SEM, Tokyo,
Japan) (
microscopy (TEM) (Philips Tecnai G220 S-TWIN, Amsterdam, The Netherlands)
(
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