Whatman no 3 filter paper

RE were collected [33 (link)]. Tomato seeds (Lycopersicon esculentum cv. Pantelosa), obtained from the Ministry of Agriculture, Egypt, were sterilized using NaOCl (sodium hypochlorite solution at 2% v/v, for 2 min) followed by washing three times in SDW (sterile distilled water). In a Petri dish, seeds (100 seeds) were put on Hoagland solution-wetted filter papers in the darkness at 25 °C for germination. Exudates from the roots were gathered in a magnate stirrer with a perforated tray filled with 80 mL of Hoagland solution. Eighty germinated seedlings with their roots in the solution were put on the tray. Exudates were obtained by filtration through Whatman No. 3 filter papers after 1 week of growth at 18 °C to eliminate solid plant contents, and then frozen quickly with liquid nitrogen. On solidified King’s B medium, a sample of exudates was taken directly from the magnate vessels and checked for contamination. The solid material was re-dissolved in 2 mL sterile water after the exudates were lyophilized. These exudates were purified through a 0.45 µm Millipore membrane to eliminate undissolved particles and deposited at −20 °C until needed. Samples were identified and confirmed using liquid chromatography-mass spectrometry (Agilent, Santa Clara, CA, USA) [34 (link)].

The samples used in this study were collected between May and August 2018 as described earlier [16 (link), 17 (link)]. Demographic data were obtained from each consenting participant, and about 2 ml of venous blood was collected into an EDTA-vacutainer® tube. Thick and thin blood smears were prepared and processed for malaria parasite identification and quantification as previously described [2 (link)]. Dried blood spots on filter paper were prepared by spotting four 50-µl drops of the venous blood onto a Whatman® No. 3 filter paper (Whatman plc, Maidstone, UK), following which the blood spots were air-dried and stored individually in zip-lock bags containing silica gels. Packed cells were then isolated from the remaining venous blood and stored in a 1.5-ml Eppendorf tube at − 20 °C for future use [16 (link)].

As published previously,^{10 (link)} 30 high-transmission and 30 low-transmission health facility catchment areas (HFCAs) were randomly assigned to one of three arms of the study: MDA, focal MDA (fMDA), and control (n = 10 HFCAs in each). Within each of the 60 total HFCAs, roughly 40 individuals older than 3 months were enrolled in a nested longitudinal cohort and surveyed monthly by community health workers for the duration of the study (18 months).^{11 (link)} At each visit, the following were ascertained: finger-prick blood samples for Giemsa microscopy (first 6 months only), an RDT (SD Bioline Malaria Ag P.f, Standard Diagnostics, Gyeonggi-do, Republic of Korea), and dried blood spot (DBS). Individuals testing positive were treated with the national standard of care. Dried blood spots were collected on Whatman 903 protein saver cards (first 6 months) or Whatman no. 3 filter paper (remainder of study) (Whatman^®, Maidstone, United Kingdom), dried during collection, packed in standard plastic “pill” bags with desiccants, and stored (< 1 month) at room temperature before being transported to the laboratory at the National Malaria Control Centre in Lusaka, Zambia, for storage at −20°C.

Sterile fungal filtrate was obtained for assessing their effects on garden cress seed germination and seedling growth according to Ogórek [5 (link)]. Five discs (0.3 cm diameter) of mycelia and spores were taken from the periphery of 10-day-old culture of each isolate grown on PDA medium and introduced into 50 mL of Sabouraud dextrose broth (peptone 10 g L⁻¹ and glucose 40 g L⁻¹) in 250-mL conical flasks. Czapek Dox broth (BioCorp) was used in the case of enzyme assay. The flasks were incubated at 25 ± 1 °C for 14 days. Cell-free fungal filtrates were obtained by passing the cultures through a sterile Whatman No. 3 filter paper followed by passage through a sterile filter (0.45 μm).

For extraction of compounds, the bacteria were cultivated in 1000-mL flasks containing 300 mL DVR1 medium (6.7 g malt extract (Sigma), 11.1 g peptone from casein, enzymatic digest (Sigma), 6.7 g yeast extract (Sigma), 0.5 L filtered sea water, and 0.5 L ddH₂O) cultures for 16 days, at 10 °C and 130 rpm. A total of 12 flasks were inoculated, giving 3.6 L of culture. The medium was autoclaved for 30 min at 120 °C prior to inoculation. Cultures were started by loop inoculation from the non-axenic glycerol stock solution.
Extraction of metabolites from the liquid media was done with Diaion^® HP-20 resin (Supelco, Bellefonte, PA, USA). The resin was activated by incubation in methanol for 30 min, followed by washing with ddH₂O for 15 min, and added to the cultures (40 g/L). The cultures were incubated with resin for 3 days prior to compound extraction. For extraction, the resin beads were separated from the liquid by vacuum filtration through a cheesecloth mesh (Dansk Hjemmeproduktion, Ejstrupholm, Denmark), the resin was washed with ddH₂O, and finally extracted two times with methanol. The extract was vacuum filtered through a Whatman No. 3 filter paper (Whatman plc, Maidstone, UK), and dried under reduced pressure at 40 °C.

Black radishes were obtained from Sungsan Ilchubong Nonghyup, whose planting facility is located in Jeju Province, Korea. Black radish roots (5000 g) were cut into thin slices, dried at 45 °C, and extracted three times at 80 °C–90 °C for 6 h with distilled water. The extracts were filtered through Whatman no. 3 filter paper and then concentrated 5-fold at 44 °C using a Buchi R-210 rotary evaporator. The BRHE was dried in a freeze-drying oven for 3 days and then stored at −20 °C.