[et_pb_section fb_built=”1″ _builder_version=”4.16″ _module_preset=”default” background_color=”#FFFFFF” custom_padding=”||0px||false|false” global_colors_info=”{}”][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif|700|||||||” text_text_color=”#4D1749″ text_font_size=”19px” global_colors_info=”{}”]<\/p>\n
With the COVID-19 pandemic still spreading across the globe, the fight is on to rapidly develop efficient solutions for diagnosis and treatment. In the past, cell-free protein expression has been used to produce antibodies against other strains of coronavirus. As such, could cell-free systems pose a viable option towards finding a solution against SARS-CoV-2 ?<\/p>\n
[\/et_pb_text][\/et_pb_column][\/et_pb_row][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n
Throughout recent years, there has been promising research using cell-free systems<\/strong> to produce antibodies for either diagnostic tests<\/strong> or neutralizing treatments<\/strong>. For coronaviruses, this has been successfully done before. In particular, previous studies have been able to produce coronavirus antigens<\/strong> and, consequently, antibodies against them [1]. Therefore, there is reason to believe that it can be done again for the current or future outbreaks.<\/p>\n The two major steps<\/strong> in producing an antibody of this type are (a) producing the target antigen<\/strong> and (b) then, generating, selecting and producing a specific antibody<\/strong> against that antigen. Both of which require expression of a highly specific protein in its native form. Hence, it must be correctly folded, without aggregation and expressed at a sufficient yield. Cell-free protein synthesis offers a reliable solution. The technique is known for its ability to produce proteins that are particularly \u201cdifficult\u201d to express in their native form using traditional in vivo systems.<\/p>\n In 2016, researchers in Japan successfully developed antibodies to precisely diagnose a strain of coronavirus known as MERS-CoV<\/strong> [1]. MERS-CoV is a virus first identified back in 2012, which causes an illness known as Middle East Respiratory Syndrome<\/strong>. Whilst the antibodies they produced would not attack the virus in COVID-19<\/strong>, their study does show how the cell-free procedure can generate very specific antibodies which can be used in the search for a solution.<\/p>\n [\/et_pb_text][\/et_pb_column][\/et_pb_row][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” header_2_font=”PT Serif|700|||||||” header_2_font_size_tablet=”” header_2_font_size_phone=”24px” header_2_font_size_last_edited=”on|desktop” global_colors_info=”{}”]<\/p>\n [\/et_pb_text][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n The first step in making antibodies is to produce an antigen. A key part of the breakthrough in the 2016 study was taking the structure of MERS-CoV into account. As with many coronaviruses, MERS-CoV is comprised of four main proteins. Most importantly, the spike (S) proteins<\/strong>, which give coronaviruses their characteristic prickly surface, are heavily targeted by host antibodies. This is thought to drive them to have a high mutation rate, so unlikely to be winners when it comes to making a specific diagnostic test or a vaccine.<\/p>\n On the other hand, the nucleocapsid (N) protein<\/strong>, which is present in high levels in infected cells, is more conserved. Since N-proteins are less likely to change over time, these antigens make better targets for specific diagnostic tests. The team therefore chose this sequence to synthesize as a protein against which they could then raise antibodies.<\/p>\n [\/et_pb_text][\/et_pb_column][\/et_pb_row][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” header_2_font=”PT Serif|700|||||||” header_2_font_size_tablet=”” header_2_font_size_phone=”24px” header_2_font_size_last_edited=”on|phone” global_colors_info=”{}”]<\/p>\n [\/et_pb_text][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n The N-protein that the team had selected is a viral structural protein, which tend to be insoluble. That means traditional systems of protein expression are often unsuccessful in producing the molecule in its native form. This is problematic. If the protein is not correctly folded<\/strong>, the antibodies produced further down the line will not detect the correct sequence.<\/p>\n In cell-free, proteins are produced in a test tube using intra-cellular machinery from cellular extract. The viruses, which are in many cases cytotoxic, cannot kill the host cell so expression rates are much higher than in vivo systems. Therefore, the authors opted for a cell-free protein expression system<\/strong>. In the end, the Japanese team successfully obtained N-protein that was not only properly folded, but also in quantities similar to what can be expected for human cells.<\/p>\n [\/et_pb_text][\/et_pb_column][\/et_pb_row][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” header_2_font=”PT Serif|700|||||||” header_2_font_size_tablet=”” header_2_font_size_phone=”24px” header_2_font_size_last_edited=”on|phone” global_colors_info=”{}”]<\/p>\n [\/et_pb_text][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n Once the protein was produced and isolated, it was injected into mice to obtain antibodies against it. The success was from the specificity of the antibody that was produced. As they were required for use in a diagnostic test, the monoclonal antibody<\/strong> binding site needed to be specific to the MERS-CoV virus<\/strong>. Their results showed that it could specifically recognize the desired N-protein, meaning that it could be used to distinguish this coronavirus from other coronaviruses.<\/p>\n Using a cell-free system the team in this study produced a high-quality native protein<\/strong>, specific enough to effectively produce antibodies. It is also worth considering that other studies have shown that, once sequenced, the antibody synthesis<\/strong> can be done using a cell-free system<\/strong>, too [2,3].<\/p>\n There is already research being carried out elsewhere at the moment using cell-free protein synthesis to produce neutralizing antibodies<\/strong> in the hopes of finding a way to efficiently fight against COVID-19 [4]. In this particular case, using a cell-free system also enabled a \u201cgame-changing\u201d rapid upscale of production<\/strong>, reducing manufacturing time from nine months to only one<\/strong> [4]. And, during a pandemic, every second counts.<\/p>\n [\/et_pb_text][\/et_pb_column][\/et_pb_row][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” custom_padding=”|15px||15px|true|true” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n Whilst we hope that the research will be fruitful, the need to raise antibodies for research or clinical purposes goes beyond that of COVID-19. Cell-free systems provide a highly promising option<\/strong> for future study. Hence, the technique could be used to produce coronavirus N-protein for a future a therapeutic solution, diagnostic tests; or even to quickly develop and produce a vaccine.<\/span><\/p>\n<\/blockquote>\n [\/et_pb_text][\/et_pb_column][\/et_pb_row][\/et_pb_section][et_pb_section fb_built=”1″ theme_builder_area=”post_content” _builder_version=”4.21.0″ _module_preset=”default”][et_pb_row _builder_version=”4.21.0″ _module_preset=”default” column_structure=”1_2,1_2″ theme_builder_area=”post_content”][et_pb_column _builder_version=”4.21.0″ _module_preset=”default” type=”1_2″ theme_builder_area=”post_content”][\/et_pb_column][et_pb_column _builder_version=”4.21.0″ _module_preset=”default” type=”1_2″ theme_builder_area=”post_content”][dnxte_button _builder_version=”4.21.0″ _module_preset=”default” theme_builder_area=”post_content” button_text=”Talk to us” button_link=”@ET-DC@eyJkeW5hbWljIjp0cnVlLCJjb250ZW50IjoicG9zdF9saW5rX3VybF9wYWdlIiwic2V0dGluZ3MiOnsicG9zdF9pZCI6IjE5ODgifX0=@” _dynamic_attributes=”button_link” background_color=”#4D1749″ btn_fonts_text_color=”#EDE8EC” dnxt_button_alignment=”right” hover_enabled=”0″ sticky_enabled=”0″][\/dnxte_button][\/et_pb_column][\/et_pb_row][\/et_pb_section][et_pb_section fb_built=”1″ _builder_version=”4.16″ _module_preset=”default” background_color=”#FFFFFF” custom_padding=”||0px||false|false” global_colors_info=”{}”][et_pb_row _builder_version=”4.16″ _module_preset=”default” width=”100%” global_colors_info=”{}”][et_pb_column type=”4_4″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][et_pb_divider color=”#D2C6D2″ _builder_version=”4.16″ _module_preset=”default” global_colors_info=”{}”][\/et_pb_divider][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” header_2_font=”PT Serif|700|||||||” header_4_font=”PT Serif|700|||||||” header_4_text_color=”#D2C6D2″ header_4_font_size=”26px” header_2_font_size_tablet=”” header_2_font_size_phone=”24px” header_2_font_size_last_edited=”on|phone” header_4_font_size_tablet=”26px” header_4_font_size_phone=”24px” header_4_font_size_last_edited=”on|phone” global_colors_info=”{}”]<\/p>\n [\/et_pb_text][et_pb_text _builder_version=”4.16″ _module_preset=”default” text_font=”PT Serif||||||||” text_text_color=”#000000″ text_font_size=”17px” text_line_height=”1.8em” global_colors_info=”{}”]<\/p>\n [1] Development of Monoclonal Antibody and Diagnostic Test for Middle East Respiratory Syndrome Coronavirus Using Cell-Free Synthesized Nucleocapsid Antigen<\/a><\/p>\nChoosing the target<\/h2>\n
Producing the antigen<\/h2>\n
Making the antibody<\/h2>\n
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Sources<\/h4>\n