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Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a...

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Autores principales: Cuttitta, Christina M., Ericson, Daniel L., Scalia, Alexander, Roessler, Christian G., Teplitsky, Ella, Joshi, Karan, Campos, Olven, Agarwal, Rakhi, Allaire, Marc, Orville, Allen M., Sweet, Robert M., Soares, Alexei S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304690/
https://www.ncbi.nlm.nih.gov/pubmed/25615864
http://dx.doi.org/10.1107/S1399004714013728
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author Cuttitta, Christina M.
Ericson, Daniel L.
Scalia, Alexander
Roessler, Christian G.
Teplitsky, Ella
Joshi, Karan
Campos, Olven
Agarwal, Rakhi
Allaire, Marc
Orville, Allen M.
Sweet, Robert M.
Soares, Alexei S.
author_facet Cuttitta, Christina M.
Ericson, Daniel L.
Scalia, Alexander
Roessler, Christian G.
Teplitsky, Ella
Joshi, Karan
Campos, Olven
Agarwal, Rakhi
Allaire, Marc
Orville, Allen M.
Sweet, Robert M.
Soares, Alexei S.
author_sort Cuttitta, Christina M.
collection PubMed
description Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s(−1)) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.
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spelling pubmed-43046902015-01-30 Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening Cuttitta, Christina M. Ericson, Daniel L. Scalia, Alexander Roessler, Christian G. Teplitsky, Ella Joshi, Karan Campos, Olven Agarwal, Rakhi Allaire, Marc Orville, Allen M. Sweet, Robert M. Soares, Alexei S. Acta Crystallogr D Biol Crystallogr New Mounts Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s(−1)) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening. International Union of Crystallography 2015-01-01 /pmc/articles/PMC4304690/ /pubmed/25615864 http://dx.doi.org/10.1107/S1399004714013728 Text en © Cuttitta et al. 2015 http://creativecommons.org/licenses/by/2.0/uk/ This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.
spellingShingle New Mounts
Cuttitta, Christina M.
Ericson, Daniel L.
Scalia, Alexander
Roessler, Christian G.
Teplitsky, Ella
Joshi, Karan
Campos, Olven
Agarwal, Rakhi
Allaire, Marc
Orville, Allen M.
Sweet, Robert M.
Soares, Alexei S.
Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title_full Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title_fullStr Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title_full_unstemmed Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title_short Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
title_sort acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening
topic New Mounts
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304690/
https://www.ncbi.nlm.nih.gov/pubmed/25615864
http://dx.doi.org/10.1107/S1399004714013728
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