Fluorescence decay of dyed protozoa: differences between stressed and non-stressed cysts
Several series of tests have shown that fresh, intact samples of Giardia duodenalis and Cryptosporidium parvum (oo)cysts are not marked by fluorescent probes such as carboxyfluorcein‐succinimidyl‐diacetate‐ester (CFDA‐SE), C12‐resazurin and SYTOX® Green, probably because of their robust cell walls....
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| Veröffentlicht in: | Luminescence (Chichester, England) England), 2015-11, Vol.30 (7), p.1139-1147 |
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| creator | Santos, Samuel Ricardo dos Branco, Nilson Franco, Regina Maura Bueno Paterniani, José Euclides Stipp Katsumata, Masakazu Barlow, Peter W. de Mello Gallep, Cristiano |
| description | Several series of tests have shown that fresh, intact samples of Giardia duodenalis and Cryptosporidium parvum (oo)cysts are not marked by fluorescent probes such as carboxyfluorcein‐succinimidyl‐diacetate‐ester (CFDA‐SE), C12‐resazurin and SYTOX® Green, probably because of their robust cell walls. These dyes fail to indicate the viability of such protozoa and allow negative responses to be recorded from living and infectious samples. Cryptosporidium parvum showed stronger isolation from chemicals, with living oocysts remaining unstained by the probe for up to 90 days after extraction. However, in further fluorescence decay (FD) experiments run with G. duodenalis samples stained using CFDA‐SE (comprising living, non‐stressed but aged cysts, heat‐killed samples and UV‐C‐stressed samples) each showed a different FD decay profile, here studied in seven series of tests of five replicates each. The FD profiles were fitted by double‐exponential decay kinetics, with the decay constant k2 being five times higher than k1. This FD procedure is fast and can be easily reproduced in 10 steps, taking ~ 1 h of laboratory work for already purified samples. Copyright © 2015 John Wiley & Sons, Ltd. |
| doi_str_mv | 10.1002/bio.2872 |
| format | Article |
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These dyes fail to indicate the viability of such protozoa and allow negative responses to be recorded from living and infectious samples. Cryptosporidium parvum showed stronger isolation from chemicals, with living oocysts remaining unstained by the probe for up to 90 days after extraction. However, in further fluorescence decay (FD) experiments run with G. duodenalis samples stained using CFDA‐SE (comprising living, non‐stressed but aged cysts, heat‐killed samples and UV‐C‐stressed samples) each showed a different FD decay profile, here studied in seven series of tests of five replicates each. The FD profiles were fitted by double‐exponential decay kinetics, with the decay constant k2 being five times higher than k1. This FD procedure is fast and can be easily reproduced in 10 steps, taking ~ 1 h of laboratory work for already purified samples. 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These dyes fail to indicate the viability of such protozoa and allow negative responses to be recorded from living and infectious samples. Cryptosporidium parvum showed stronger isolation from chemicals, with living oocysts remaining unstained by the probe for up to 90 days after extraction. However, in further fluorescence decay (FD) experiments run with G. duodenalis samples stained using CFDA‐SE (comprising living, non‐stressed but aged cysts, heat‐killed samples and UV‐C‐stressed samples) each showed a different FD decay profile, here studied in seven series of tests of five replicates each. The FD profiles were fitted by double‐exponential decay kinetics, with the decay constant k2 being five times higher than k1. This FD procedure is fast and can be easily reproduced in 10 steps, taking ~ 1 h of laboratory work for already purified samples. Copyright © 2015 John Wiley & Sons, Ltd.</description><subject>carboxyfluorescein-diacetate-succinimidyl-ester</subject><subject>Coloring Agents - chemistry</subject><subject>Constants</subject><subject>Cryptosporidium parvum</subject><subject>Cryptosporidium parvum - chemistry</subject><subject>Cysts</subject><subject>Cysts - chemistry</subject><subject>Decomposition</subject><subject>dyes</subject><subject>Fluorescence</subject><subject>fluorescence decay</subject><subject>Giardia duodenalis</subject><subject>Giardia lamblia</subject><subject>Giardia lamblia - chemistry</subject><subject>Luminescence</subject><subject>Protozoa</subject><subject>Walls</subject><issn>1522-7235</issn><issn>1522-7243</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqN0VFL3TAYBuAwJtOpsF8wCrvZTTXJ16Tp7jadTnSKoLi7kCZfoK6n0aRFu1-_HDw7gjDwKiE8ecmXl5APjO4xSvl-24U9rmr-hmwxwXlZ8wrervcgNsn7lG4ppVLK5h3Z5EI2FKDaIr-O-ilETBYHi4VDa-Yi-MLN6Iq7GMbwJ5gvheu8x7gkqWhxfEAcijTmaykzM7hiCEO5PrBzGtMO2fCmT7i7WrfJ9dH3q4Mf5dnF8cnB17PSCg68bD0FRq1ngMrxVlLmHBe1AeVspYxA5NhKZF55YYwSsq2ow7ZxSKFSFmGbfH7Kza-9nzCNetHlafreDBimpFldU8ixkr6CVlIpxsRrKBf5c6uGZfrpBb0NUxzyzEtVMQBQzXOgjSGliF7fxW5h4qwZ1csOde5QLzvM9OMqcGoX6NbwX2kZlE_goetx_m-Q_nZysQpc-S6N-Lj2Jv7WsoZa6JvzY32jGFwe_jzVp_AXG02zuQ</recordid><startdate>201511</startdate><enddate>201511</enddate><creator>Santos, Samuel Ricardo dos</creator><creator>Branco, Nilson</creator><creator>Franco, Regina Maura Bueno</creator><creator>Paterniani, José Euclides Stipp</creator><creator>Katsumata, Masakazu</creator><creator>Barlow, Peter W.</creator><creator>de Mello Gallep, Cristiano</creator><general>Blackwell Publishing Ltd</general><general>Wiley Subscription Services, Inc</general><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QF</scope><scope>7QO</scope><scope>7QP</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7U5</scope><scope>7U7</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F1W</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H95</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L.G</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope><scope>M7N</scope></search><sort><creationdate>201511</creationdate><title>Fluorescence decay of dyed protozoa: differences between stressed and non-stressed cysts</title><author>Santos, Samuel Ricardo dos ; 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These dyes fail to indicate the viability of such protozoa and allow negative responses to be recorded from living and infectious samples. Cryptosporidium parvum showed stronger isolation from chemicals, with living oocysts remaining unstained by the probe for up to 90 days after extraction. However, in further fluorescence decay (FD) experiments run with G. duodenalis samples stained using CFDA‐SE (comprising living, non‐stressed but aged cysts, heat‐killed samples and UV‐C‐stressed samples) each showed a different FD decay profile, here studied in seven series of tests of five replicates each. The FD profiles were fitted by double‐exponential decay kinetics, with the decay constant k2 being five times higher than k1. This FD procedure is fast and can be easily reproduced in 10 steps, taking ~ 1 h of laboratory work for already purified samples. Copyright © 2015 John Wiley & Sons, Ltd.</abstract><cop>England</cop><pub>Blackwell Publishing Ltd</pub><pmid>25690334</pmid><doi>10.1002/bio.2872</doi><tpages>9</tpages></addata></record> |
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| ispartof | Luminescence (Chichester, England), 2015-11, Vol.30 (7), p.1139-1147 |
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| source | MEDLINE; Wiley Online Library Journals Frontfile Complete |
| subjects | carboxyfluorescein-diacetate-succinimidyl-ester Coloring Agents - chemistry Constants Cryptosporidium parvum Cryptosporidium parvum - chemistry Cysts Cysts - chemistry Decomposition dyes Fluorescence fluorescence decay Giardia duodenalis Giardia lamblia Giardia lamblia - chemistry Luminescence Protozoa Walls |
| title | Fluorescence decay of dyed protozoa: differences between stressed and non-stressed cysts |
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