Lab6 tem

Scanning electron microscopy (SEM) was performed using a Hitachi S-4800 field emission microscope at an accelerating voltage of 10 kV. The samples were prepared on Si wafers, then sputtered with a very thin layer of Au/Pd. BET surface area measurements were performed with a Nova 2200e, Quantachrome Instruments. FT-IR was performed on a Perkin-Elmer SpectrumBX instrument fitted a SensIR Technologies DuraSampleIR II ATR unit. Transmission electron microscopy (TEM) was performed on a 200 kV JEOL LaB6 TEM. The in situ video was captured using a Hysitron PI95 TEM Picoindenter accessory, Hysitron Corporation, Minneapolis, MN.

After 42 days of feeding, one fish from each tank was randomly euthanized with AQUI-S at 175 mg/L to excise liver, muscle, gill, and intestine for histological and TEM evaluation in response to test diets. For histological analysis, samples of all organs were fixed immediately in 10% buffered formalin, subsequently dehydrated with series of ethanol, infiltrated in xylene, and embedded in paraffin wax, as per standard histological protocols. Section of approximately 5 μm thickness was stained with Periodic Acid-Schiff (PAS) and digitally photographed under a light microscope (BX40F4, Olympus, Tokyo, Japan).
For TEM analysis, freshly collected intestinal samples washed in 2.5% glutaraldehyde buffered in 1x PBS at pH 7.4 before performing secondary fixation in 1% OsO4 (80 W 2 min on, 2 min off, 2 min on), dehydrating in ethanol (50, 70, 95 and 100% at 250 W, 40 seach) and infiltrating finally with epoxy resin in acetone (Procure 812, Proscitech) (1:3, 1:1, 3:1ratios at 250 W, 3 min each). Samples were processed as described in the earlier study in our lab [16 (link)] and screened a LaB6 TEM (JEOL2100, Japan) at 120 kV. The electron micrographs obtained from TEM analysis at 30,000 magnification were analysed using ImageJ (National Institute of Health, USA) to determine microvilli length and diameter.