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"Indonesian National Committee for human immunodeficiency virus (HIV) and acquired immune deficiency syndrome(AIDS) has reported the significant increase of HIV infected individual in Indonesia. A sensitive accurate diagnosticsare urgently needed to prevent the dissemination of HIV and also to provide a suitable therapy. For this reason, we havedeveloped HIV diagnostic method based on PCR to elucidate this problem. For this research, samples were collectedfrom hospitals and Indonesian Red Cross that consist of samples possesing HIV serological test positive andindeterminate. Ribonucleic Acid (RNA) were isolated from blood plasma. These RNA then were amplified afterReverse transcriptase reaction by using primers which have been designed using env (C2V3C3), Long TerminalRepeats (LTR) (U3) and Capsid gag (p24) as target regions. The obtained results shown 26/34 (76.5%) positive in LTR,10/33 (36.4%) positive in Env and 11/33 (33.3%) positive in p24. These results showed that LTR primers detect HIVmore than other primers. It suggests that LTR could be used as detection target as complement of env and p24 Theseresults need to be explored further by using sequencing method."

"Beberapa hasil uji serologi HIV indeterminate pada tes skrining darah ditemukan di Indonesia. Prosedur skrining darahyang dilakukan saat ini sesuai ketentuan yang ditetapkan oleh WHO untuk skrining darah, yaitu 3 tes uji serologi HIVselama pemeriksaan darah. Ketidaksesuaian hasil yang satu dengan yang lain didefinisikan sebagai hasil indeterminate.Penelitian ini bertujuan untuk mengidentifikasi galur-galur HIV yang sulit teridentifikasi dari darah dengan uji serologiHIV intermediate dan mengevaluasi apakah galur HIV yang beredar di Indonesia mempunyai kemungkinan lolos darisistem pendeteksian yang ada. Deteksi RT-PCR dilakukan pada 40 sampel RNA HIV dari donor darah yangmempunyai hasil uji serologi indeterminate dengan sebelumnya melakukan uji konfirmasi dengan menggunakanwestern blot. Deteksi RT-PCR menunjukkan bahwa sebanyak 24/32 (75%) sampel positif LTR, 4/31 (13%) positif poldan 3/5 (60%) positif env. Amplifikasi pada daerah p24, pita-pita yang ditemukan pada sampel selalu lebih rendah dariyang diharapkan. Sekuensing dilakukan untuk mengkonfirmasi hasil amplifikasi menunjukkan bahwa perlu analisislebih lanjut untuk mengetahui apakah perubahan ini yang menyebabkan hasil indeterminate.Indeterminate results ofserological HIV test have been found in Indonesia. The screening procedure is following the prescribed by WHO forscreening of blood donors which is based on 3 different serological HIV test during screening of blood donors.Discordant results are interpreted as indeterminate. This research aims to identify GIV strains that previously difficult todetermine, and to evaluate whether the HIV strains present in Indonesia could pass the existing screening system. RTPCRdetection test of HIV RNA were conducted for 40 blood donors samples with indeterminate serological HIV-testafter a confirmatory test using western blot. Preliminary results showed that 24/32 (75%) of the samples are positiveLTR, 4/31 (12%) positive pol and 1/3 (33%) positive env. Amplification in p24 region showed that bands found havelower size than expected. Sequencing performed to confirm these findings show that further analysis is needed todetermine whether this change is what behind the indeterminate results."

"Protein Transaktivator transkripsi (Tat) adalah protein regulator HIV-1 berfungsi sebagai aktivator transkripsi genom HIV-1. Varian protein Tat-Eli adalah aktivator transkripsi paling kuat daripada varian lain melalui induksi promotor LTR HVI-1. Kemampuan tersebut digunakan sebagai kontrol positif dalam pengembangan uji infeksitivitas HIV-1 berbasis gene reporter eGFP diregulasi LTR HIV-1. Pengembangan uji infeksivitas tersebut menawarkan waktu deteksi infeksi lebih singkat daripada uji p24 pada uji fenotipik. Penelitian ini bertujuan untuk mengekspresikan protein rekombinan Tat-Eli di sistem ekpresi prokariot dan mempurifikasinya sehingga dapat dijadikan kontrol positif penginduksi promotor. Pada penelitian ini dilakukan pengklonaan gen sintetik Tat-Eli ke vektor pQE80L. Protein rekombinan Tat-Eli dipurifikasi menggunakan Ni-NTA. Pengklonaan ulang gen reporter eGFP disisipkan setelah promotor LTR HIV-1. Aktivitas protein rekombinan Tat-Eli terhadap ekspresi eGFP di sel mamalia dinilai berdasarkan persentase sel pengekspresi eGFP dan intensitas cahaya eGFP.Konstruksi plasmid rekombinan membawa gen Tat-Eli, pQETat, berhasil dibuat, diekspresikan dan dipurifikasi kondisi native. Plasmid pengekspresi eGFP  dengan promoter HIV-1, pLTReGFP berhasil dikonstruksi. Penambahan Tat-Eli rekombinan pada sel mamalia yang ditransfeksi pLTReGFP menunjukkan perbedaan intensitas cahaya eGFP yang bermakna dan paling tinggi dari semua perlakuan. Protein rekombinan Tat-Eli dapat diekspresikan dan dipurifikasi secara optimal dari E.coli. Penambahan protein Tat-Eli pada sel yang ditransfeksi pLTReGFP meningkatkan intensistas cahaya eGFP.

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Transcriptional Transactivator Protein (Tat) is an HIV-1 regulatory protein functioning as an activator of HIV-1 genome transcription. The Tat-Eli protein variant was the most potent transcriptional activator than other variants through the induction of the HVI-1 LTR promoter. This ability was used as a positive control in the development of an HIV-1 infection test based on the eGFP reporter gene regulated by LTR HIV-1. The development of the infectiousness test offers a shorter infection detection time than the p24 test in the phenotypic test. This study aims to express Tat-Eli recombinant protein in the prokaryotic expression system and to purify it so that it can be used as a positive control inducer of the promoter. In this study, synthetic Tat-Eli gene was cloned into the pQE80L vector. Tat-Eli recombinant protein was purified using Ni-NTA. Recloning of the eGFP reporter gene was inserted after the HIV-1 LTR promoter. The activity of Tat-Eli recombinant protein on eGFP expression in mammalian cells was assessed based on the percentage of eGFP-expressing cells and eGFP light intensity. The recombinant plasmid construction carrying the Tat-Eli gene, pQETat was successfully generated, expressed and purified in native conditions. An eGFP-expressing plasmid with HIV-1 promoter, pLTReGFP was successfully constructed. The addition of recombinant Tat-Eli to mammalian cells transfected with pLTReGFP showed a significant difference in eGFP light intensity and was the highest of all treatments. Tat-Eli recombinant protein can be optimally expressed and purified from E. coli. The addition of Tat-Eli protein in pLTReGFP-transfected cells increased eGFP light intensity."