Abstract
Large number of second-stage juveniles of root-knot nematode, Meloidogyne incognita was prepared for scanning electron microscopy over a millipore filter (pore size 0.45 µm). The J2 juveniles were quickly immobilized and killed before fixation using the cryo-fixation technique followed by the primary fixation in 2.5% glutaraldehyde pre-cooled at 4-8°C and then secondary vapor fixation in 1% osmium tetraoxide and drying with the series of ethanol without subsequent critical point drying. The specimens were vacuum dried in a desiccator for a period of 24-48 h. Cryo-fixation emphasizes the need to quickly stop cellular activity and thus prevent any surface artifacts that occur due to conventional fixation techniques. Key words: Scanning electron microscopy, cryo-fixation, root-knot nematode, Meloidogyne incognita, surface morphology.
Highlights
Technique fo or en-m masse cryo-ffixation n and proces ssing o of seco ond-sta age juv veniles s of Me eloidog gyne in ncogn nita for scanni s ng electron m micros scopy
Proce essing technique was eva aluated on the e basis of the e final iimage obtaine ed with the sscanning electron micrroscope (SEM)
The quallity of presserrvation n of the J2 su urface using the cryo-fixattion technique e and o obtaining the em in large n number using g the millipore e filter p paper technique were the m most important observations seen with the SEM
Summary
Large number of secon nd-stage juv veniles of root-knot nem matode, Melo oidogyne inc cognita was prepared for scannin ng electron microscopy over a milllipore filter C. Cryo-fixation n emphasize es the need tto quickly stop cellula ar activity and a pre event any su urface artifa acts that occ cur due to c conventiona al fixation techniques. Meloidogyne inc matode considered as one e of the most damaging crrop nem pessts affecting crop c productio on Specimen n prepa aration remains a limiting g factor in tthe quality of o inform mation provided by SEM. SEM investiga ations of plant-pa arasitic nema atodes comp pared severa al differe ent methods of specimen preparation
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