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Journal : Indonesian Journal of Biotechnology

Combination PT-PCR with in vitro Transcription-Translation System: Rapid and Simple Approach for Monoclonal Antibody Production Ali, Muhamad
Indonesian Journal of Biotechnology Vol 14, No 2 (2009)
Publisher : Universitas Gadjah Mada

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Abstract

We have developed a simple and efficient system for screening and generation of monoclonal antibodies, which can bypass conventional monoclonal antibody production system that includes time-consuming, labor-intensive establishment and cultivation. Our method consist of: (1) cDNA synthesis from single B cells of immunized mouse, followed by (ii) PCR amplification of the Lc and Hc (Fd portion) cDNAs separately using low concentration (0,05 ìM) of the respective cDNA-specific primers with 5’ homotags in the presence of the homotag-specific primer (0,5 ìM), (iii) overlapping PCR of the amplified Lc and Hc cDNAs with the cassettes for the T7 promoter (T7P) and T7 terminator (T7T) to construct the following expression units: T7P-Lc-T7T and T7P-Hc-T7T, and (iv) in vitro expression of these units using an Escherichia coli S30 extract followed by ELISA screening without</div><div>purification. Light-and heavy-chain cDNA were amplified and expressed using in vitro system, thus avoiding time consuming steps of cloning and bacterial expression. Having successfully amplified and expressed Lc and Hc genes from single B cells in rapid, simple, versatile, labor-saving, and cost effective, this method seem to be useful</div><div>and applicable for the high-throughput monoclonal antibody generation. 
Co-Authors . Adwia . Eliya, . . Juliana . Jumini . Mira, . . NURHAYATI . Nurkholifah, . . Setiawati, . A.A. Ketut Agung Cahyawan W Abdi, Aurell Vinaya Abdullah, Sirajudin H. Adang Kuswaya Agniny, Siswi Fililmi Ahmad Taufik S Aidillah Suja Ainunnissa, Shereen Regina Akhmad Taufiq Mukti Al Gifari, Zaid Al'Aqsa, Muhammad Ramadhan Alawiyah, Geby Alfina Taurida Alaydrus Ali Ridho Alorinda Maruli, Alorinda Amanda, Shafira Putri Amelina, Rosa Aminurrahman Amudin, Amudin Ana, Nurul Anjasmara, L. Dede Arta Satya Annisa Amalia Anton Anton Anwar Rosyidi Aprianti, Rika Aprilia Novitasari Ardiana Ekawanti Aryana F64112029) Asih Priyati Aswandi . Atlanta, Tatiek Aulia, Syarifah Baiq Rismarini Nursaly, Baiq Rismarini Busri Endang Dahlanuddin Dahlanuddin, Dahlanuddin Danang Prio Utomo Depamede, Sulaiman N Desni Yuniarn, Desni Desni Yuniarni Dian Miranda Dian Utami Dewi Dinamella Wahjuningrum Djoko Laras Budiyo Taruno Dwi Khairunnisyah, Dwi Dwi Retnowati Dwiyanti, Devi Dzikri, Ardhika Ekawati (F54209006) Elina, Ernamitha Erawati, Yeni Eugenie, Olivia Esty Fadila, Salima Fadillah . Fahreza, Firdha Khriska Fahrudin Fahrudin Fajriyah, Putri Arifatul Farhana Muhammad Faruq, Rafif Umar Fatimah, Dayang Srilia Firmansah, Firmansah Fitriyan Fadjar Suryadi, Muh. Aidil Flora, Elina Fransiska RZ FUKUBA, Takako Gifari, Zaid Al Gita Deviana, Gita Gusti Jhoni Putra, Gusti Jhoni Gustiana, Della Hadi, L. Asnan Kurnia Al Halida . Halida Halida Halil Akhyar Handoko, Heldi Hanifah . Harianto, Biko Hasibuan, Zanniro Sururi Hasma Hasma Henny Noviyeni Hery Heryanto Hidayatullah, Kholik himmawan, Didik Husaefa, Nadira Huzain Jailani ica, Stevania IFA, KHOIRUL Iis Suyantini Ikhwan Firhamsah Ima Arum Lestarini Imron Hadi Indri Astuti Indrianasari, Neny Tri Ine Karni Irsyad, Akhmad Irtia, Fitri Jamhari Jamhari Jayadi, Iguk Jessica, Agustina Juani, Muhamad Jumiatin, Nur Junia Ayu Ninggrum, ⁠Vidia Khairil Anwar Khairunnisa, Alya Syafira Khurrotul Akyunin Kiki Aryanita, Kiki Koprita, Yusela Kurnia, F. Devi Kurniati F54209020 Kurniawidi, Dian W. Laili Mardiana Lalu Muhammad Fauzi Lalu Parhanuddin Lalu Sahrul Hudha Leni Dahlia Lidiastuti, Nazila Lilis Sumarni Linarsih, Andini Linda Yulianti Lukmanulhakim ., Lukmanulhakim Lusina, Lusina M TAUFIQ M. Syukri M. Syukri . M. Thamrin Made Sriasih, Made Maharani, Rini Mahasari, Gunanti Mardiyanti Mardiyanti Marhaini, Marhaini Marianty, . Marisa Dwi Sukmawati, Marisa Dwi Mariska, Tina Vidya Marmawi R . Marudin Mas ud, Aan Masluyah Suib Mawazi, Abd Rahman Merry, Adriana Meydiawanti, Salsabilla Meyzawinata, Nabil Abdillah Miranti Eka Pratiwi MIRNAWATI SUDARWANTO Morsek, Wita Kristina Mudinillah, Adam Muh. Fahrurrozi Muhamad Amin Muhamad Juaini MUHAMMAD ANSHORI, MUHAMMAD Muhammad Yusuf Muhammad Zainul Majdi Muhfizaturrahmah, Muhfizaturrahmah Muis, Muhamad Ali Murad, Murad Najmi, Aulia NAKANO, Hideo Narsi (F54010014) Nasution, Raeni Sariyanti Nayyara, Najma Silma Neni Endang Kusniarti Nila Hayati Ningsih, Sri Rahayu Nisa, Faridhatun Nony Amelia Novialita, Jessica Noviana, Desi Nurdiah . Nurhening Yuniarti Nurlinayati ., Nurlinayati Nurmaulida, Siti Ema Nuro Sholihah Nurul Hasanah Nurul Yuliasari Oczalina, Bella Olifia, Nonita Pratmaningsih, Sri Astuti PURWANTI PURWANTI Puteri, Sylvana Putra, Ryan Aryadin RA Maheswari, Rarah Rahayu, Umi Puji Rahmat, Syahrul Ratnasari, Erma Restiana, . Resty, Polina Rezki Amalyadi Rif’atul Khoiriah Malik Rina Andriati, Rina Rita, Helena Rody Satriawan Rosyidi, Muhammad Subhan Bahruddin Rotin, Rotin Rozak, Rama Wijaya Abdul Rukmana, Hajar Safitri, Risa Nur Sahrul Alim Salawati, Suchi Saleha, Risti Saputra, Zira Sarjan, M Selvianita, Selvianita Septian, I Gede Nano Septiana, Riska Setyadi, Hario Jati Sholihah, Isfi Sidik, Muhammad Alfan Sinta, Sinta Siti Salmah Slamet Riyadi Sochib Sochib, Sochib Sri Wahyuni Subhani, Armin - Sukarne Sukarne Sulaiman Ngongu Depamede Sulasmi, Neni Sunarti Suriati, Ira Suryadi, Muh. Aodil Fitriyan Fadjar Susi Rahayu Susilawati, Susilawati Susiowati, Susiowati Sutarmanto . Tanuadi, I Tegi, Lidia Krisdayanti Thibburruhany, Thibburruhany Toto Sukisno Ulfa, Vizza Novian Unsunnidhal, Lalu Veronika Leni, Veronika Wahyu Ilaihi Wahyuni, Yuni Wariata, Wayan Widyadhana, Talita Ardra Wintia, Sisilia Lilik Wirawan, Rahadi Yazid, Muh Yuliana Yuliana Yuline . Yuniarti, Desni Yunita Sabrina, Yunita Yuny Erwanto Zaid Al Gifari Zain, Rohaeniyah Zainal Arifin Zariah, . Zein, Rohaeniyah Zulfina, Septi