1997
DOI: 10.1046/j.1365-2818.1977.2390796.x
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Freeze shattering: a simple and effective method for permeabilizing higher plant cell walls

Abstract: SummaryThis article describes a practical technique for permeabilization of higher plant cell walls, which is usually one of the first steps required for immunolocalization of cellular components (and other cytological methods) in plant cell studies. Our strategy involves shattering the walls of cells while the tissues are frozen in liquid nitrogen. It replaces the use of wall degrading enzymes or the need to employ laborious sectioning or other mechanical means for providing access of probes to cells. Freeze-… Show more

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Cited by 97 publications
(66 citation statements)
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“…Microtubule localization to the convex regions of cells has been reported for lobed pavement cells for many species (Panteris and Galatis, 2005) and has been reported repeatedly in Arabidopsis pavement cells (Wasteneys et al, 1997;Fu et al, 2002Fu et al, , 2005Qiu et al, 2002;Zhang et al, 2011;Sampathkumar et al, 2014). Our TEM analyses here detected a similar bias for microtubule localization to the convex regions of cells ( Fig.…”
Section: Discussionsupporting
confidence: 87%
See 1 more Smart Citation
“…Microtubule localization to the convex regions of cells has been reported for lobed pavement cells for many species (Panteris and Galatis, 2005) and has been reported repeatedly in Arabidopsis pavement cells (Wasteneys et al, 1997;Fu et al, 2002Fu et al, , 2005Qiu et al, 2002;Zhang et al, 2011;Sampathkumar et al, 2014). Our TEM analyses here detected a similar bias for microtubule localization to the convex regions of cells ( Fig.…”
Section: Discussionsupporting
confidence: 87%
“…Subsequently, the pavement cells expand for days and increase in area largely independent of lobe initiation (Zhang et al, 2011;Armour et al, 2015;Wu et al, 2016). In highly lobed pavement cells, anticlinal microtubules often are concentrated at the convex region of the cell (Wasteneys et al, 1997;Zhang et al, 2011), and this microtubule arrangement may reflect the ability of the microtubule network to maintain polarized lobe outgrowth and rearrange in response to local cell geometry and the predicted patterns of cell wall stress (Sampathkumar et al, 2014). Transverse microtubules within expanding lobes (concave cell domain) also may pattern microfibrils to reinforce highly polarized lobe outgrowth.…”
mentioning
confidence: 99%
“…Transformed Arabidopsis seedlings were fixed, permeabilized by freeze-shattering in liquid nitrogen, and immunolabeled as described previously (Wasteneys et al, 1997). Double immunolabeling of membrane-bound GFP and endoplasmic reticulum (ER)-specific BiP epitopes was achieved by combining two mouse GFP-specific monoclonal antibodies (each at a working dilution of 1:200; Boehringer Mannheim, Mannheim, Germany; Clontech, Palo Alto, CA) with the rabbit anti-BiP antibodies used in immunoblotting experiments (1:1000 dilution).…”
Section: Immunofluorescence Techniquesmentioning
confidence: 99%
“…with Figures 4H to 4O). The sporadic permeabilization of single epidermal cells by the freeze-shatter method (Wasteneys et al, 1997) provided a convenient way to contrast antibody-specific fluorescence with GFP fluorescence. The upper guard cells shown in Figures 6C and 6D, for example, were not permeabilized and consequently display only residual GFP fluorescence, whereas the permeabilized lower guard cells (and pavement cell shown in Figure 6B) are brighter because the GFP has been labeled with FITC.…”
Section: Membrane-bound Gfp Colocalizes With Bipmentioning
confidence: 99%
“…Van Lammeren et al 1985;Hause et al 1993). Because cells and tissues are three-dimensional objects, and the spatial organisation of their components are of importance for their function, whole mount immunolocalisation is preferred and can be achieved by enzymatic digestion of the cell wall (Lloyd et al 1979) or freeze shattering (Wasteneys et al 1997). The whole mount approach is particularly useful for visualizing structural elements such as microtubules and actin filaments, which is the aim of this article.…”
Section: Introductionmentioning
confidence: 99%