Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA)
Serum bactericidal assay (SBA) is the method to investigate in vitro complement-mediated bactericidal activity of sera raised upon vaccination. The assay is based on incubating the target bacteria and exogenous complement with sera at different dilutions and the result of the assay is represented by...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-09-01
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| Series: | BioTech |
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| Online Access: | https://www.mdpi.com/2673-6284/10/3/19 |
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| author | Maria Grazia Aruta Martina Carducci Francesca Micoli Francesca Necchi Omar Rossi |
| author_facet | Maria Grazia Aruta Martina Carducci Francesca Micoli Francesca Necchi Omar Rossi |
| author_sort | Maria Grazia Aruta |
| collection | DOAJ |
| description | Serum bactericidal assay (SBA) is the method to investigate in vitro complement-mediated bactericidal activity of sera raised upon vaccination. The assay is based on incubating the target bacteria and exogenous complement with sera at different dilutions and the result of the assay is represented by the sera dilution being able to kill 50% of bacteria present in the inoculum. The traditional readout of the assay is based on measurement of colony-forming units (CFU) obtained after plating different reaction mixes on agar. This readout is at low throughput and time consuming, even when automated counting is used. We previously described a novel assay with a luminescence readout (L-SBA) based on measurement of ATP released by live bacteria, which allowed to substantially increase the throughput as well as to reduce the time necessary to perform the assay when compared to traditional methods. Here we present a further improvement of the assay by moving from a 96-well to a 384-well format, which allowed us to further increase the throughput and substantially reduce costs while maintaining the high performance of the previously described L-SBA method. The method has been successfully applied to a variety of different pathogens. |
| first_indexed | 2024-03-10T07:51:45Z |
| format | Article |
| id | doaj.art-e67d0f7de42d46a4bd21e1fc38191ab0 |
| institution | Directory Open Access Journal |
| issn | 2673-6284 |
| language | English |
| last_indexed | 2024-03-10T07:51:45Z |
| publishDate | 2021-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | BioTech |
| spelling | doaj.art-e67d0f7de42d46a4bd21e1fc38191ab02023-11-22T12:13:09ZengMDPI AGBioTech2673-62842021-09-011031910.3390/biotech10030019Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA)Maria Grazia Aruta0Martina Carducci1Francesca Micoli2Francesca Necchi3Omar Rossi4GSK Vaccines Institute for Global Health (GVGH) S.r.l., 53100 Siena, ItalyGSK Vaccines Institute for Global Health (GVGH) S.r.l., 53100 Siena, ItalyGSK Vaccines Institute for Global Health (GVGH) S.r.l., 53100 Siena, ItalyGSK Vaccines Institute for Global Health (GVGH) S.r.l., 53100 Siena, ItalyGSK Vaccines Institute for Global Health (GVGH) S.r.l., 53100 Siena, ItalySerum bactericidal assay (SBA) is the method to investigate in vitro complement-mediated bactericidal activity of sera raised upon vaccination. The assay is based on incubating the target bacteria and exogenous complement with sera at different dilutions and the result of the assay is represented by the sera dilution being able to kill 50% of bacteria present in the inoculum. The traditional readout of the assay is based on measurement of colony-forming units (CFU) obtained after plating different reaction mixes on agar. This readout is at low throughput and time consuming, even when automated counting is used. We previously described a novel assay with a luminescence readout (L-SBA) based on measurement of ATP released by live bacteria, which allowed to substantially increase the throughput as well as to reduce the time necessary to perform the assay when compared to traditional methods. Here we present a further improvement of the assay by moving from a 96-well to a 384-well format, which allowed us to further increase the throughput and substantially reduce costs while maintaining the high performance of the previously described L-SBA method. The method has been successfully applied to a variety of different pathogens.https://www.mdpi.com/2673-6284/10/3/19serum bactericidal assayvaccinefunctional assayhigh throughputluminescent SBA |
| spellingShingle | Maria Grazia Aruta Martina Carducci Francesca Micoli Francesca Necchi Omar Rossi Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) BioTech serum bactericidal assay vaccine functional assay high throughput luminescent SBA |
| title | Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) |
| title_full | Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) |
| title_fullStr | Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) |
| title_full_unstemmed | Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) |
| title_short | Increasing the High Throughput of a Luminescence-Based Serum Bactericidal Assay (L-SBA) |
| title_sort | increasing the high throughput of a luminescence based serum bactericidal assay l sba |
| topic | serum bactericidal assay vaccine functional assay high throughput luminescent SBA |
| url | https://www.mdpi.com/2673-6284/10/3/19 |
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