Didik T. Subekti
Balai Besar Penelitian Veteriner, JL. RE Martadinata 30, Bogor 16114, Indonesia

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Parasitaemia pattern and mortality of mice infected by Indonesian Isolate of Trypanosoma evansi. DH, Sawitri; Subekti, Didik T.; AH, Wardhana; ., Suhardono
Indonesian Journal of Animal and Veterinary Sciences Vol 18, No 4 (2013)
Publisher : Indonesian Animal Sciences Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (2410.495 KB) | DOI: 10.14334/jitv.v18i4.334

Abstract

Trypanosomiasis (Surra) is one of the parasitic diseases is endemic and deadly for horses and buffalo in Indonesia.The etiology of the disease is a Trypanosoma evansi. Some T. evansi isolates had been isolated and cryopreservated. Those isolates had not been studied for their differences in virulence, particularly with regard to the pattern of parasiteaemia and their ability to promote mice mortality. Therefore in this study the differences in virulence was studied. DDY mice were divided in to 19 groups according to each isolate to be tested. Each group consisted of 5 mice. Infection were carried intraperitoneally at a dose of 104 Trypanosoma/mice. Mice were examined every two days. Blood samples were taken from tail’s peripheral blood and were examined under light microscope. Parasite were quantitatively counted using Naubauer chamber.  Parasitemia and mice survival were observed for 30 days or until all mice died.The results indicated that there was significant difference among the isolates.Through out the nineteenth isolate scan could be grouped into 3 different biotypes associated with patterns of parasitemia and their ability to kill mice. Biotype1 was the most virulent with the ability to promote mice mortality ≤ 8 days post-infection (dpi). The biotype2 and 3 were the lowest compared to biotype 1. Biotype2 had an undulating parasitaemia, where as biotype 3 showed persistently high parasitaemia with the ability to promote mice mortality ≥ 14dpi. The results also indicate the presence of mixed infections of biotypes that exist in one isolate of T. evansi. Key Words: Trypanosoma evansi, Biotype, Virulence, Mixed infection, Parasitaemia
Comparison of serological test for toxoplasmosis using Immunostick Assay, ELISA and Latex Agglutination Test Subekti, Didik T.; Kusumaningtyas, E.
Indonesian Journal of Animal and Veterinary Sciences Vol 16, No 3 (2011)
Publisher : Indonesian Animal Sciences Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (233.755 KB) | DOI: 10.14334/jitv.v16i3.617

Abstract

Many serological methods were developed in order to improve detection and identification of an infectious diseases. ELISA (enzyme linked immunosorbent assay), is a well-known as the most serological technique and widely used for infectious disease diagnoses, especialy toxoplasmosis in human and animal. Unfortunately, the technique requires specific tools and expertise that are not always available in the field. Inn addition, ELISA needs longer process than other techniques for both small and large number of samples. Another serological assay, namely Latex Agglutination Test (LAT) was known as populer and mostly applied in field for toxoplasmosis diagnosis in Indonesia, both for human and animal serum. However, the accuracy of this technique was slightly lower compared to ELISA, especially for weakly seropositive. Recently, an immunostick assay technique was developed as alternative serological method which has high accuracy as ELISA. The immunostick assay was rapid and simple like latex agglutination test. The immunostick assay was evaluated to detect Toxoplasma gondii infection in goat serum samples and compared to ELISA and LAT. The immunostick assay had a true agreement 95.88% - 100% againts ELISA and their strength of agreement was very good aggreement with ELISA (k = 0.796; AC1 = 0.948). However, LAT had true agreement only 75.74%, either with ELISA or immunostick assay. The strength of agreement of LAT was moderately aggreement with ELISA or immunostick assay (k = 0.447; AC1 = 0.582). Therefore, the immunostick assay could become the first choice for rapid serological assay on toxoplasmosis diagnosis, both in the laboratory and in the field. Key Words: Latex Agglutination, ELISA, Immunostick Assay, FELISA, Toxoplasmosis
Perbandingan Penggunaan Konjugat Antibovine IgG-HRP dan Protein A/G-HRP Dengan Beberapa Larutan Pengencer Serum Pada ELISA Untuk Deteksi Surra pada Sapi dan Kerbau Subekti, Didik T.; Yuniarto, Ichwan
JURNAL BIOLOGI INDONESIA Vol 13, No 1 (2017): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (822.292 KB) | DOI: 10.14203/jbi.v13i1.3093

Abstract

ABSTRACTSurra was reported in various types of animals both livestock, pets and wildlife. ELISA (enzyme linked immunosorbentassay) is a sensitive and specific diagnosis technique for Surra detection. The use of speciesspecific conjugate (anti bovine IgG-HRP) has limited while protein A/G can be used for a wide variety of animalspecies. This study aimed to evaluate the initial application of the protein A/G-HRP compared with antibovine IgG-HRP using standard samples using four (4) types of diluent buffer. The standard serum samples (23serum) consist of positive and negative sera from bovine (cattle and buffalo) were reacted with Surra antigenon microplate. Positive and negative serum was diluted with different diluent buffer, namely PBS-Tween20(PBST), RBA (RedBuff A), RBB (RedBuff B) and LC (Low Cross). Results of ELISA using protein A/G-HRPshowed absorbance values reduced 36.16% - 69.30% compared to the anti-bovine IgG-HRP. The percentagereduction of PBST, RBA, RBB and LC, were 51.76%; 56.64%; 36.16% and 69.30% respectively. The use ofprotein A/G-HRP and fourth diluent buffer can reduce antigen - antibody bonding with a weak affinity whichlowers the absorbance value of ELISA Surra.Keywords : Protein A, Protein G, Surra, ELISA, Trypanosoma evansi
Performa Perangkat Diagnostik Elisa Toksoplasmosis pada Serum Domba dan Manusia Subekti, Didik T.; Hayati, Lisda; Raharja, Sujud M.
JURNAL BIOLOGI INDONESIA Vol 8, No 2 (2012): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14203/jbi.v8i2.3052

Abstract

Performance of ELISA Toxoplasmosis Diagnostic Kits on Sheep and Human Sera Toxoplasma seropositivity in Indonesia have a high prevalence, both in human and animals. Unfortunately, the availability of diagnostic tools to support dynamic surveillance are limited. Recently, the diagnostic tools for toxoplasmosis, namely ELISA BM were developed. The technology was based on ELISA technique using soluble tachyzoite antigen from tachyzoites of Toxoplasma gondii. Kit performance is one of the important issue for acceptance of diagnostic tools prior to wide application. The purpose of the studies was to asses the quality of diagnostic toolsperformances. The assesment comprises of four stages. First stage was to evaluate the performance of ELISA BM compared to Latex Agglutination Test (LAT) on sheep sera. Secondly, to evaluate the performance of ELISA BM to descriminate true seropositive andseronegative toxoplasmosis on human sera. The last stage were comparing ELISA BM, ELISA TL (commercial kit) and LAT on predetermined and unknown human sera. The results showthat the accuracy of ELISA BM is slightly better than ELISA TL. Agreement of ELISA BM with LAT was better againts ELISA TL with LAT. However, all performance as determined using Cohen’s ? and Gwet’s AC1 of ELISA BM, ELISA TL and LAT were good up to very good agreement.Keywords: Toxoplasmosis, ELISA, Latex Agglutination, inter reliability agreement.
Identifikasi Protein dan Seleksi Isolat Trypanosoma evansi Bersifat Imunogenik untuk Kandidat Pengembangan Imunoasai Subekti, Didik T.; Yuniarto, Ichwan
JURNAL BIOLOGI INDONESIA Vol 14, No 1 (2018): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (1180.87 KB) | DOI: 10.14203/jbi.v14i1.3669

Abstract

ABSTRACTSurra is a parasitic disease caused by T. evansi and causing high economic losses in Indonesia. Some isolates have been isolated from several areas experiencing outbreaks of Surra in Indonesia. The isolates have been reported to have a diversity of protein profiles based on SDS PAGE (sodium dodecyl sulphate polyacrilamide gel electrophoresis). The research aims to identify immunogenic protein from each isolate and make the selection of T. evansi isolates which is potential as a source of antigens for immunoassay development. Each isolates were obtained to be purified from blood and then the protein was isolated. The proteins were run onto polyacrylamide gel electrophoresis and visualized by Coomassie blue. Another electrophoresis results were transferred onto a nitrocellulose membrane for immunoblotting. The results showed that the immunogenic proteins that consistently detected among nine T. evansi isolates of Indonesia are 100, 90, 85, 76-80, 70, 65, 55, 49-52, 44-46, 40, 34-36, 31 -33 kDa. Among the nine T. evansi isolates, N372 isolate was selected as a candidate for immunoasay development, especially ELISA. Immunogenic proteins were specifically found on the N372 isolate are 85, 70, 65, 49-52, 44-46, 34-36, 31-33, 24-28, 15-20 kDa.Keywords : Trypanosoma evansi, immunoblotting, protein profiles, immunogenic protein
PERFORMA PERANGKAT DIAGNOSTIK ELISA TOKSOPLASMOSIS PADA SERUM DOMBA DAN MANUSIA Subekti, Didik T.; Hayati, Lisda; Raharja, Sujud M.
JURNAL BIOLOGI INDONESIA Vol 8, No 2 (2012): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14203/jbi.v8i2.3052

Abstract

Performance of ELISA Toxoplasmosis Diagnostic Kits on Sheep and Human Sera Toxoplasma seropositivity in Indonesia have a high prevalence, both in human and animals. Unfortunately, the availability of diagnostic tools to support dynamic surveillance are limited. Recently, the diagnostic tools for toxoplasmosis, namely ELISA BM were developed. The technology was based on ELISA technique using soluble tachyzoite antigen from tachyzoites of Toxoplasma gondii. Kit performance is one of the important issue for acceptance of diagnostic tools prior to wide application. The purpose of the studies was to asses the quality of diagnostic toolsperformances. The assesment comprises of four stages. First stage was to evaluate the performance of ELISA BM compared to Latex Agglutination Test (LAT) on sheep sera. Secondly, to evaluate the performance of ELISA BM to descriminate true seropositive andseronegative toxoplasmosis on human sera. The last stage were comparing ELISA BM, ELISA TL (commercial kit) and LAT on predetermined and unknown human sera. The results showthat the accuracy of ELISA BM is slightly better than ELISA TL. Agreement of ELISA BM with LAT was better againts ELISA TL with LAT. However, all performance as determined using Cohen?s ? and Gwet?s AC1 of ELISA BM, ELISA TL and LAT were good up to very good agreement.Keywords: Toxoplasmosis, ELISA, Latex Agglutination, inter reliability agreement.
IDENTIFIKASI PROTEIN DAN SELEKSI ISOLAT TRYPANOSOMA EVANSI BERSIFAT IMUNOGENIK UNTUK KANDIDAT PENGEMBANGAN IMUNOASAI Subekti, Didik T.; Yuniarto, Ichwan
JURNAL BIOLOGI INDONESIA Vol 14, No 1 (2018): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14203/jbi.v14i1.3669

Abstract

ABSTRACTSurra is a parasitic disease caused by T. evansi and causing high economic losses in Indonesia. Some isolates have been isolated from several areas experiencing outbreaks of Surra in Indonesia. The isolates have been reported to have a diversity of protein profiles based on SDS PAGE (sodium dodecyl sulphate polyacrilamide gel electrophoresis). The research aims to identify immunogenic protein from each isolate and make the selection of T. evansi isolates which is potential as a source of antigens for immunoassay development. Each isolates were obtained to be purified from blood and then the protein was isolated. The proteins were run onto polyacrylamide gel electrophoresis and visualized by Coomassie blue. Another electrophoresis results were transferred onto a nitrocellulose membrane for immunoblotting. The results showed that the immunogenic proteins that consistently detected among nine T. evansi isolates of Indonesia are 100, 90, 85, 76-80, 70, 65, 55, 49-52, 44-46, 40, 34-36, 31 -33 kDa. Among the nine T. evansi isolates, N372 isolate was selected as a candidate for immunoasay development, especially ELISA. Immunogenic proteins were specifically found on the N372 isolate are 85, 70, 65, 49-52, 44-46, 34-36, 31-33, 24-28, 15-20 kDa.Keywords : Trypanosoma evansi, immunoblotting, protein profiles, immunogenic protein
PERBANDINGAN PENGGUNAAN KONJUGAT ANTIBOVINE IGG-HRP DAN PROTEIN A/G-HRP DENGAN BEBERAPA LARUTAN PENGENCER SERUM PADA ELISA UNTUK DETEKSI SURRA PADA SAPI DAN KERBAU Subekti, Didik T.; Yuniarto, Ichwan
JURNAL BIOLOGI INDONESIA Vol 13, No 1 (2017): JURNAL BIOLOGI INDONESIA
Publisher : Perhimpunan Biologi Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14203/jbi.v13i1.3093

Abstract

ABSTRACTSurra was reported in various types of animals both livestock, pets and wildlife. ELISA (enzyme linked immunosorbentassay) is a sensitive and specific diagnosis technique for Surra detection. The use of speciesspecific conjugate (anti bovine IgG-HRP) has limited while protein A/G can be used for a wide variety of animalspecies. This study aimed to evaluate the initial application of the protein A/G-HRP compared with antibovine IgG-HRP using standard samples using four (4) types of diluent buffer. The standard serum samples (23serum) consist of positive and negative sera from bovine (cattle and buffalo) were reacted with Surra antigenon microplate. Positive and negative serum was diluted with different diluent buffer, namely PBS-Tween20(PBST), RBA (RedBuff A), RBB (RedBuff B) and LC (Low Cross). Results of ELISA using protein A/G-HRPshowed absorbance values reduced 36.16% - 69.30% compared to the anti-bovine IgG-HRP. The percentagereduction of PBST, RBA, RBB and LC, were 51.76%; 56.64%; 36.16% and 69.30% respectively. The use ofprotein A/G-HRP and fourth diluent buffer can reduce antigen - antibody bonding with a weak affinity whichlowers the absorbance value of ELISA Surra.Keywords : Protein A, Protein G, Surra, ELISA, Trypanosoma evansi