Smz1270 stereo microscope

Colony-forming units (CFUs) in semi-solid methylcellulose were quantified to assess hematopoietic progenitor activity. WBM was flushed from femurs and tibiae using a 26G x 1/2 needle with MACS buffer. Viable cell counts were determined with a hemocytometer using Trypan Blue. WBM cells were plated in duplicate in Methocult GF M3434 methylcellulose (StemCell Technologies 03444) according to the manufacturer’s recommendations. Colonies were scored for phenotypic CFU-GEMM, CFU-GM, CFU-G, CFU-M, and BFU-E colonies using a SMZ1270 Stereo-Microscope (Nikon).

For immunofluorescence, spleens were embedded and frozen in OCT, sectioned at 15 μm and mounted on SuperFrostPlus Adhesion glass (Thermo Fisher Scientific). Sections were dehydrated using silica beads, fixed with 4% paraformaldehyde for 10 min and washed with PBS. Samples were blocked using 5% normal goat serum for 2 h before staining. Samples were incubated with antibodies against B220 (RA3-6B2, eBioscience), CD3 (17A2, eBioscience) and F4/80 (BM8, Biolegend) diluted in 5% NGS for 2 h at room temperature in the dark. After staining, samples were washed with PBS at least three times. Samples were then mounted using ProLong Gold Antifade Mountant (Invitrogen) and imaged using an inverted LSM780 microscope (Carl Zeiss) and a plan apochromat 63× NA 1.40 oil-immersion objective (Carl Zeiss). For haematoxylin and eosin (H&E) staining, organs were collected and fixed in 10% formalin. Fixed samples were embedded in paraffin and sectioned at 10 μm, mounted on SuperFrostPlus Adhesion glass and stained using H&E. Mounted samples were imaged using a Nikon SMZ1270 Stereo Microscope. Imaging data were analysed using Fiji (ImageJ) software (NIH).

Studies were conducted on 1- or 2-day-old final-instar larvae of European corn borers (fifth instar), corn earworms (sixth instar), fall armyworms (sixth instar), painted ladies (fifth instar), red admirals (fifth instar), and wax moths (sixth instar). All larvae were placed in individual rearing containers and randomly assigned a treatment prior to starting a bioassay. One μL of acetone or an imidacloprid-acetone solution was placed on the dorsal prothorax using a 10 μL pipette. Five imidacloprid concentrations and an acetone control were used. At least ten larvae were treated in each of the six groups and five control larvae were weighed prior to treatment. Daily observations for mortality were taken until pupation and at adult eclosion. Signs of larval intoxication (e.g., spasms, paralysis) and formation of malformed pupae were recorded. Dissections were performed on a subset of larvae that had AE. Photographs were taken with a Nikon DS-Ri2 digital camera connected to a Nikon SMZ1270 stereo microscope. The results obtained were compared to topical imidacloprid exposures conducted on fifth instar monarch butterfly^{14 (link),15 (link)} to provide a more complete description of AE.

H₂O₂ was detected in roots using the 2',7'-dichlorodihydrofluorescein diacetate (H₂DCFDA) staining method. The roots from 7-day-old seedlings were incubated in H₂O containing 50 µM H₂DCFDA for 30 min under dark conditions. Then, the roots were rinsed with sterilized water three times. The fluorescent signals were observed with an Echo Revolution microscope (Echo). The excitation and emission wavelengths used for detection of the signals were 488 nm and 520 nm, respectively.
To detect superoxide, seedlings were stained for 2 min in a solution of 2.5 mM Nitroblue tetrazolium (NBT) in 50 mM phosphate buffer (pH 6.1) in the dark and then rinsed three times with distilled water^{49 (link)}. Images for NBT staining were obtained using a 1× objective with a Nikon SMZ1270 Stereo microscope.

Pictures of biofilm pellicles of GA1 formed at the air-liquid interface were taken with a Nikon SMZ1270 stereomicroscope (Nikon, Japan) equipped with a Nikon DS-Qi2 monochrome microscope camera and a DS-F 1× F-mount adapter. Pictures were captured in the bright field channel with an ED Plan Apo 1×/WF objective and a 0.63x magnification of with an exposure time of 40ms. Additionally, pictures of biofilm pellicles at 6h were captured with a 3x magnification and an exposure time of 10ms to get a better view of the structures in formation. Acquisitions were then processed with NIS-Element AR software (Nikon, Japan).