Nakisah Mat Amin* and ¹Keith Vickerman

Unit of Biological Sciences, Faculty of Science and Professional Arts, Universiti Putra Malaysia Terengganu, Mengabang Telipot 21030 Kuala Terengganu, Malaysia, 1Division of Environmental Evolutionary Biology, Graham Kerr Building, University of Glasgow, G12 8QQ, Scotland, UK.

(Received 9 January 1998 / Accepted 23 February 1998)

Abstract.
A fluorescence technique has been employed to visualise nuclear division in Naegleria gruberi. The synchronously dividing Naegleria cells were stained with anti a-tubulin as a primay antibody and with a fluorescein isothiocyanate labelled goat anti mouse immunoglobin (FITC) as a secondary antibody. The cells were also stained with 4 1 6-diamidino-2-phenylindole DAP1) and propidium iodide (PI) to stain the nucleus. The results of this study indicated that by using anti (a tubulin, cytoplasmic microtubules; and mitotic spindles of Naegleria gruberi can be visualised under fluorescence microscopy. By this technique, a centrosome with associated microtubules was found to be present in the cytoplasm of Naegleria during nuclear division and this finding contradicts the previous idea that this organelle does not exist in Naegleria. The centrosome appeared to divide during prophase but it could not be detected later at metaphase telophase.

Keywords: Anti a tubulin, 4', 6 diamidino 2 phenylindole DAP1) Propidium Iodide (PI), Centrosome, Spindle Microtubules

Y,F. Ngeow*, L Rajan, V, Hema and CA Gaydos¹

Department of Medical Microbiology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.
The Johns Hopkins University, Chlamydia Research Laboratory, Baltimore, USA.

(Received 29 December 1998 / Accepted 10 January 1998)

Abstract.
A major outer membrane protein (MOMP) based nested polymerase chain reaction (PCR) for the detection of Chlamydia trochomatios was evaluated for urine specimens collected from patients attending sexually trans. mitted disease clinic. A hundred and thirty nine samples were tooted by the nested VCR and a commercially available ligase chain reaction (LCR) ;assay called the LCx from Abbott Laboratories, USA, To resolve discrepant results, 87 of these specimen were also tested by a second LCR amplifying it di (OMPI) gene than the plasmi.d gene targetted in the LCx, The remainder 32 specimens were examined by the IDEIA Chlamydia, an enzyme immunoassay from DAKO Diagnostic@ Ltd,, UK, Using an expanded "gold standard" consisting of two positive test results, the sensitivity, specificity, positive find negative predictive values of the nested VCR were found to be 55.1%, 98,0%, 98,0% and 55.1% respectively. There was no significant difference in the test porformance for male and female urine

Keywords: Chlamydia Trachomatis, Urine, PCR

Wan Omar Abdullah*, Low Kiat Cheong¹, Vivian Jacob Gomes¹, Sulaiman Osman¹ and Yusof Suboh¹

Department of Medical Sciences, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor;
¹Department of Parasitology and Medical Entomology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur

(Received 27 October 1997 / Accepted 16 June 1998)

Abstract.
The application of dispersed dye particles as label in immunological assay was investigated as an alternative to enzyme labeled immunoassay (ELISA). Dipsticks prepared from the antibody (Ab) coated nitrocellulose membrane mounted on acetate strips served as capture matrix. Ab absorbed to a commercial brilliant pink colloidal dye particles served as antigen (Ag) detecting reagent. The dipstick developed was compared with plate ELISA for detecting antigenemic cases of Dirofilaria immitis infection in dogs. Evaluation statistics of the dipstick assay varied according to the cutoff value of the ELISA titers. The overall specificity and sensitivity of ELISA was 96.67% and 98.57% respectively. The dipstick performed with a specificity of 85.71 % and a sensitivity of 94.12% when the ELISA titre was 160. There was high level of agreement between these two assay formats ranging from 89.47% to 92.60%. Dipstick assay detected 1% of the occult dirofiliariasis as with the plate ELISA. As the dipstick assay is simpler, rapid, inexpensive, equally sensitive and specific its use in detecting active cases of D. immitis infections in dogs is recommeded especially under field condition. The dipstick immunoassay format would enable veterinary practitioners to quickly diagnose and appropriately treat most cases of the disease.

Keywords: colourimetric, dipstick, inununoassay, dirofilariasis

Garcia, R., Pimentel, E., Somonte, D., Mena, J., Zaldua, Z., Lopez, A., Moran, R., and Garcia*, M.

Centro de lngenieria Genetica y Biotecnologia. P.O. Box 387, C.P. 70100. Camaguey, CUBA

(Received 2 February 1998 / Accepted 16 July 1998)

Abstract.
Isolation of sweet potato protoplasts from different organs has been very difficult to develop, due to the recalcitrant behavior of this crop when it is manipulated in-vitro. An efficient protocol for protoplast isolation from leaf and stem tissues was established. Different enzyme combinations and concentrations were tested, as well as several digestion conditions. A hard enzymatic treatment, combined with mechanical separation of the cells from the digested tissue, led to a high protoplast yield. Other factors, such as the osmotic solution used in the different washing steps, and the starch concentration in the medium, were evaluated for their influence on the final protoplast recovery and viability. The transformation of isolated protoplasts was mediated by polyethylene glycol (PEG). The plasmid transient expression was detected by a spectrophotometrical P glucoronidase (GUS) assay.

Keywords: sweet potato, protoplast isolation, PEG mediated transformation