{"id":7674,"date":"2014-01-13T09:23:00","date_gmt":"2014-01-13T17:23:00","guid":{"rendered":"http:\/\/mlmlblog.wordpress.com\/?p=7674"},"modified":"2020-10-08T13:02:44","modified_gmt":"2020-10-08T20:02:44","slug":"ballast-water-and-epifluorescence-microscopy","status":"publish","type":"post","link":"https:\/\/mlml.sjsu.edu\/student-life\/2014\/01\/13\/ballast-water-and-epifluorescence-microscopy\/","title":{"rendered":"Ballast water and epifluorescence microscopy"},"content":{"rendered":"
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by Liz Lam, Biological Oceanography Lab<\/h3>\n
\"The<\/a>
The Golden Bear Facility, home to MLML's ballast treatment testing team<\/figcaption><\/figure>\n

Ballast water treatment and testing is a big focus here in the Biological Oceanography lab, and this is no exception even when it comes to class projects. \u00a0Last semester, I started a project aiming to improve one of our counting techniques. \u00a0I\u2019d previously written<\/a> about IMO\u2019s restriction to 10 organisms per 1,000 liters of discharged ballast water and counting zooplankton under a microscope in order to check for these results. \u00a0But when it comes to even smaller organisms, such as algae and other even tinier phytoplankton, different methods are called for.<\/p>\n

We already have a pretty clever way of quantifying such microscopic organisms by using a few chemical and optical tricks. \u00a0The first key ingredient is fluorescein diacetate, or FDA. \u00a0One of the special features of this molecule is that it can only be cleaved by certain proteins in live cells. \u00a0Once FDA is split, what remains is fluorescein, a compound that glows bright green when excited under blue light. We can then use an epifluorescence microscope to both shine the right wavelength of light and magnify a sample in order to count any green organisms. \u00a0If it glows green, then it means it\u2019s alive! \u00a0This allows us to quantify the number of live organisms that are extremely small and difficult to see.<\/p>\n

\"Epifluorescence<\/a>
Epifluorescence microscope and image capture software<\/figcaption><\/figure>\n

Unfortunately, even such a clever method has a few key disadvantages. \u00a0First of all, these water samples must be counted in a 3D well-plate, making it very difficult to find organisms at different depths. \u00a0This is like trying to count chickpeas in an Olympic sized swimming pool! \u00a0Secondly, fluorescein eventually leaks out of cells, so these samples have to be counted immediately after they\u2019ve been treated and they can\u2019t be preserved over time. \u00a0That\u2019s a bit too time-consuming and inconvenient for ballast treatment testers.<\/p>\n

\"An<\/a>
An algal culture glowing green with fluorescein under an epifluorescence microscope<\/figcaption><\/figure>\n

What I investigated at the end of last semester is the possibility of preparing samples on flat slides. \u00a0This would eliminate the depth-of-focus issue and as a bonus, allow us to take photographs of known volumes of samples. \u00a0I also experimented with a variety of fixation methods, or ways of preserving the fluorescein inside cells so that it would stay there for an extended period of time. \u00a0Surprisingly, microwaving the slides seemed to do a fairly good job of keeping the fluorescence within the cells. \u00a0These findings have given me an exciting jumping-off point for this semester!<\/p>\n

\n<\/div>\n<\/div><\/div><\/div><\/div><\/div>","protected":false},"excerpt":{"rendered":"

by Liz Lam, Biological Oceanography Lab Ballast water treatment and testing is a big focus here in the Biological Oceanography lab, and this is no exception even when it comes to class projects. \u00a0Last semester, I started a project aiming to improve one of our counting techniques. \u00a0I\u2019d previously written about IMO\u2019s restriction to 10 […]<\/p>\n","protected":false},"author":291,"featured_media":7678,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[15],"tags":[59],"class_list":["post-7674","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-drop-inblog","tag-biological-oceanography-lab"],"distributor_meta":false,"distributor_terms":false,"distributor_media":false,"distributor_original_site_name":"MLML Student Life","distributor_original_site_url":"https:\/\/mlml.sjsu.edu\/student-life","push-errors":false,"_links":{"self":[{"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/posts\/7674","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/users\/291"}],"replies":[{"embeddable":true,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/comments?post=7674"}],"version-history":[{"count":1,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/posts\/7674\/revisions"}],"predecessor-version":[{"id":20670,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/posts\/7674\/revisions\/20670"}],"wp:featuredmedia":[{"embeddable":true,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/media\/7678"}],"wp:attachment":[{"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/media?parent=7674"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/categories?post=7674"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/mlml.sjsu.edu\/student-life\/wp-json\/wp\/v2\/tags?post=7674"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}