Cam3.II K13WT, Cam3.II K13R539T (RF967), and Cam3.II K13C580Y are laboratory-adapted Cambodian isolates that were genetically engineered at the K13 locus as described in the literature 14 (link). Cam3.II K13C580Y parasites expressing β2 C31Y, β2 C31F, or β5 A20S were obtained from in vitro selection studies with WLL and WLW as described in the literature 36 (link). Parasite cultures were propagated in O+ red blood cells (RBCs) (anonymous donors, purchased from Interstate Blood Bank, Memphis, Tennessee). RBCs were stored at 50% hematocrit in ADSOL (2 mM adenine (Alfa Aesar, Haverhill, Massachusetts), 111 mM dextrose (Fisher BioReagents, Pittsburgh, Pennsylvania), 41.2 mM mannitol (Acros Organics, Fair Lawn, New Jersey), and 154 mM sodium chloride (Fisher BioReagents) for prolonged RBC vitality as described in ref 47 (link) at 4oC. Parasites were cultured in complete media (RPMI 1640 media supplemented with 0.01 mg/mL gentamicin (Gibco, Dun Laoghaire, Co Dublin, Ireland), 50 mg/mL hypoxanthine (Acros Organics), and 0.5% Albumax II (Invitrogen, Carlsbad, California)) at 5% hematocrit, and were maintained at 37oC in a Heracell™ VIOS 160i CO2 Incubator (Thermo Fisher Scientific, Waltham, Massachusetts) under hypoxic conditions (5% O2, 5% CO2, 90% N2). Gas was purchased from Matheson Gas (Irving, Texas).
Mannitol
Manufactured by Thermo Fisher Scientific
Sourced in United Kingdom, Palestine, State of
Mannitol is a polyol compound commonly used as a laboratory reagent. It serves as a cryoprotectant, stabilizer, and osmotic agent in various experimental and analytical applications.
Automatically generated - may contain errors
Lab products found in correlation
Sodium chloride (nacl), by Thermo Fisher Scientific (5 mentions)
Sucrose, by Merck Group (4 mentions)
D glucose, by Thermo Fisher Scientific (3 mentions)
Adsol, by Thermo Fisher Scientific (2 mentions)
Dextrose, by Thermo Fisher Scientific (2 mentions)
Xylose, by Thermo Fisher Scientific (2 mentions)
Galactose, by Thermo Fisher Scientific (2 mentions)
Fructose, by Thermo Fisher Scientific (2 mentions)
Urea, by Thermo Fisher Scientific (2 mentions)
Eudragit s100, by Thermo Fisher Scientific (2 mentions)
4 m naoh, by Thermo Fisher Scientific (2 mentions)
Trehalose, by Glentham Life Sciences (2 mentions)
Leucine, by Glentham Life Sciences (2 mentions)
Labplant spray dryer, by Thermo Fisher Scientific (2 mentions)
Eudragit s100, by Evonik (2 mentions)
Sorbitol, by Thermo Fisher Scientific (2 mentions)
Sucrose, by Thermo Fisher Scientific (2 mentions)
Nh4 2so4, by Merck Group (2 mentions)
Water immersion objective, by Olympus (2 mentions)
Glass capillary tube, by A-M Systems (2 mentions)
24 protocols using mannitol
1
Cultivation of Genetically-Modified Cambodian Malaria Parasites
2
Purified Native Wheat Starch Interactions
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Aytex® P wheat starch, an unmodified, highly purified native wheat starch (<0.2% protein, <0.1% fat, <0.2% ash, 9.9% water, and 25% amylose) [42 ] was donated by ADM (Minneapolis, MN, USA) and used “as is”. Twenty different sugars and sugar alcohols that may be found in food products and/or have stereochemical structures of interest were used: xylose, ribose, glucose, galactose, fructose, mannose, and mannitol from Acros Organics (Fair Lawn, NJ, USA); L-sorbose and xylitol from Sigma-Aldrich (St. Louis, MO, USA); trehalose dihydrate from Hayashibara Company (Okayama, JP, USA); tagatose and allulose from Sensato (Albany, NY, USA); maltose monohydrate and lactose monohydrate from Fisher Chemical (Fair Lawn, NJ, USA); isomaltulose monohydrate and isomalt ST (~1:1 ratio of glucopyranosyl sorbitol and glucopyranosyl mannitol dihydrate [43 ]) from BENEO-Palatinit Gmbh (Mannheim, DE, USA); sorbitol from Amresco (Solon, OH, USA); sucrose from Mallinckrodt Chemicals (Phillipsburg, NJ, USA); and maltitol and raffinose pentahydrate from Alfa Aesar (Ward Hill, MA, USA) (Table 1 ). Calcium propionate was from Sigma-Aldrich. The water used in this study was processed using reverse osmosis, then filtered by a Barnstead E-Pure Lab Water System (Dubuque, IA, USA) to >17.4 milliohm-cm.
3
Leukocyte-Depleted RBC Isolation
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O+ blood from anonymous donors was purchased from Interstate Blood Bank (Memphis, TN). Thus, no information about sex, gender, or other demographics regarding blood source is available. Whole blood was passed through a leukocyte-reduction filtration unit (Haemonetics, Braintree, MA). Then, leukocyte-depleted blood was centrifuged at 4000 rpm (3100 × g) for 10 min without brake. The supernatant was removed, and to the packed blood, an equal volume of incomplete media, which consists of RPMI 1640 media supplemented with 2.05 mM L-Glutamine (GE Healthcare, Chicago, IL), was added. This centrifugation and addition of incomplete media was performed twice to remove any residual serum, fats, white blood cells, or other components of whole blood. Pure RBCs were then resuspended at 50% hematocrit in ADSOL (2 mM adenine (Alfa Aesar, Haverhill, MA), 111 mM dextrose (Fisher BioReagents, Pittsburgh, PA), 41.2 mM mannitol (Acros Organics, Fair Lawn, NJ), and 154 mM sodium chloride (Fisher BioReagents)63 (link) for RBC preservation and stored at 4°C.
4
Radiolabeled Compound Characterization Protocol
5
Microbial Metabolism Evaluation Protocol
6
Spray-Drying Phage-Loaded Excipient Powder
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For the spray drying procedure previously described, methods were used with slight modifications (29 (link), 35 (link)). Initially, excipients and phages at titers of 5 × 1010 PFU/mL (1% phage added to final volume) were dissolved in ultrapure water to make up a 200 mL volume solution. The excipients tested were trehalose (Glentham Life Sciences Ltd, UK), leucine (Glentham Life Sciences Ltd, UK), mannitol (Fisher Scientific, UK) and Eudragit S100 (Evonik, Germany). Eudragit S100 was dissolved in ultrapurified water (5% wt/vol) by the addition of 4 M NaOH (Fisher Scientific, UK) drop by drop, until the solution turned clear, which indicated polymer dissolution (26 (link)).
The excipient-phage solution was spray dried using a laboratory scale LabPlant Spray Dryer (UK) with a two-fluid nozzle for atomization, and the nozzle had a diameter of 0.5 mm (Fig. 1a ). Air speed of 4.3 ms−1 and liquid flow rate of 280 mlh−1 were used. The drying temperature was controlled by changing the inlet temperature from 80°C to 100°C, and the corresponding outlet temperatures varied from 40°C to 60°C. Dried powder phages were passed through the cyclone, collected in 100 mL glass bottles, and stored at 4°C until use. To determine phage titer, 0.10 g of dried powder phage was suspended in 900 μL phage suspension buffer, diluted 10-fold, and titered by plaque assays (Kropinski et al., 2009).
The excipient-phage solution was spray dried using a laboratory scale LabPlant Spray Dryer (UK) with a two-fluid nozzle for atomization, and the nozzle had a diameter of 0.5 mm (
7
Microbial Growth Media and Nitrogen Sources
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Culture media (Brain heart infusion (BHI), M17, MRS, tryptic soy broth (TSB) and LB media) and inorganic nitrogen sources ((NH4)2SO4, NH6PO4, NH4NO3 and NH4Cl) were purchased from Merck (Darmstadt, Germany). Carbon sources (fructose, glucose, galactose, sucrose, lactose, maltose, sorbitol and mannitol) were obtained from Fisher Chemical (Loughborough, United Kingdom), while organic nitrogen sources (yeast extract, meat extract, peptone and soytone) were from BD (Franklin Lakes, NJ, USA).
8
Formulation of Inhalable CSP7 Trifluoroacetate Powder
9
Electrophysiological Recordings of Magnocellular Neurosecretory Cells
10
Reagent Preparation for Biochemical Assays
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Chemicals were obtained from
the following manufacturers: citric
acid, phenylalanine, sodium citrate, sucrose, deuterium oxide (D2O) 99.9% (Sigma-Aldrich Co., Gillingham, UK), mannitol, NaCl
(Fisher Scientific Inc., Loughborough, UK), and TCEP (Thermo Fisher,
Hemel Hempstead, UK).
the following manufacturers: citric
acid, phenylalanine, sodium citrate, sucrose, deuterium oxide (D2O) 99.9% (Sigma-Aldrich Co., Gillingham, UK), mannitol, NaCl
(Fisher Scientific Inc., Loughborough, UK), and TCEP (Thermo Fisher,
Hemel Hempstead, UK).
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