title: Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets.
abstract: Recently, a new technique was developed for non-catalytically mixing microdroplets. In this method, an alternating-current (AC) electric field is used to promote the antigen-antibody reaction within the microdroplet. Previously, this technique has only been applied to histological examinations of flat structures, such as surgical specimens. In this study, we applied this technique for the first time to immunofluorescence staining of three-dimensional structures, specifically, mammalian eggs. We diluted an antibody against microtubules from 1:1,000 to 1:16,000, and compared the chromatic degree and extent of fading across dilutions. In addition, we varied the frequency of AC electric-field mixing from 5 Hz to 46 Hz and evaluated the effect on microtubule staining. Microtubules were more strongly stained after AC electric-field mixing for only 5 minutes, even when the concentration of primary antibody was 10 times lower than in conventional methods. AC electric-field mixing also alleviated microtubule fading. At all frequencies tested, AC electric-field mixing resulted in stronger microtubule staining than in controls. There was no clear difference in a microtubule staining between frequencies. These results suggest that the novel method could reduce antibody consumption and shorten immunofluorescence staining time.
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Hiromitsu
authors.initials: H
authors.lastname: Shirasawa
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Jin
authors.initials: J
authors.lastname: Kumagai
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Emiko
authors.initials: E
authors.lastname: Sato
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Katsuya
authors.initials: K
authors.lastname: Kabashima
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Yukiyo
authors.initials: Y
authors.lastname: Kumazawa
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Sato
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Miura
authors.affiliation: Akita Industrial Technology Center, Akita, Japan.
authors.forename: Ryuta
authors.initials: R
authors.lastname: Nakamura
authors.affiliation: Division of Clinical Pathology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nanjo
authors.affiliation: Department of Thoracic Surgery, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Minamiya
authors.affiliation: Akita Industrial Technology Center, Akita, Japan.
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Akagami
authors.affiliation: Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine, Akita, Japan.
authors.forename: Yukihiro
authors.initials: Y
authors.lastname: Terada
mesh_terms: D000818:Animals; D004531:Eggs; D005455:Fluorescent Antibody Technique; D008322:Mammals; D051379:Mice; D008856:Microscopy, Fluorescence; D013194:Staining and Labeling