Limits...
Effects of surface passivation on gliding motility assays.

Maloney A, Herskowitz LJ, Koch SJ - PLoS ONE (2011)

Bottom Line: Beta casein did not support motility very well and averaged speeds of 870±30 nm/s.Kappa casein supported motility very poorly and we were unable to obtain an average speed.Finally, we observed that mixing alpha, beta, and kappa casein with the proportions found in bovine whole casein supported motility and averaged speeds of 966±6 nm/s.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy and Center for High Technology Materials, University of New Mexico, Albuquerque, New Mexico, United States of America. amaloney@unm.edu

ABSTRACT
In this study, we report differences in the observed gliding speed of microtubules dependent on the choice of bovine casein used as a surface passivator. We observed differences in both speed and support of microtubules in each of the assays. Whole casein, comprised of α(s1), α(s2), β, and κ casein, supported motility and averaged speeds of 966±7 nm/s. Alpha casein can be purchased as a combination of α(s1) and α(s2) and supported gliding motility and average speeds of 949±4 nm/s. Beta casein did not support motility very well and averaged speeds of 870±30 nm/s. Kappa casein supported motility very poorly and we were unable to obtain an average speed. Finally, we observed that mixing alpha, beta, and kappa casein with the proportions found in bovine whole casein supported motility and averaged speeds of 966±6 nm/s.

Show MeSH
Relative microtubule length distributions.Filled blue bars are length calculations for the alpha casein                            passivation, filled green bars are for the beta casein passivation,                            filled red bars are for the kappa casein passivation, unfilled black                            bars are for the mixed casein passivation, and unfilled purple bars are                            for the whole casein passivation. Length measurements were performed                            only on tracked microtubules and are estimations from computing the                            Convex Hull perimeter on eroded images of microtubules.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3108588&req=5

pone-0019522-g002: Relative microtubule length distributions.Filled blue bars are length calculations for the alpha casein passivation, filled green bars are for the beta casein passivation, filled red bars are for the kappa casein passivation, unfilled black bars are for the mixed casein passivation, and unfilled purple bars are for the whole casein passivation. Length measurements were performed only on tracked microtubules and are estimations from computing the Convex Hull perimeter on eroded images of microtubules.

Mentions: Figure 2 shows a histogram of microtubule lengths. To obtain lengths, we used an erosion algorithm on binary images of only the microtubules that were tracked. We used a standard function in LabVIEW/Vision 7.1 called Skeleton L. After the erosion, we used a Convex Hull perimeter calculation also found in the LabVIEW/Vision 7.1 library of functions. We divided the Convex Hull perimeter by 2 to estimate the microtubule length. Two of the authors independently and manually estimated the length of several microtubules and found that usually the Convex Hull perimeter was in between both manually obtained values. We decided that this was sufficient for identifying large changes in MT length distributions, and so we did not further investigate the robustness or systematic errors in our length estimation. As can be seen in the figure, alpha casein (filled blue bars), whole casein (empty purple line bars) and mixed casein (empty black line bars) all were able to sustain motility with smaller microtubules 2–6 µm range. Beta casein (filled green bars) had significantly fewer microtubules that were tracked, however, those that were tended to be longer than the ones tracked in the alpha casein assay. Kappa casein (filled red bars) show that there were many more trackable microtubules than beta casein and significantly fewer than alpha casein. The smallest trackable microtubules using our algorithm are not smaller than 2 µm. Figure 2 shows that while in comparison to the other lengths, there are relatively few 2 µm microtubules in any of the assays, kappa casein had no microtubules that fell in the 2–3 µm range. There is, however, a larger number of kappa casein microtubules that were in the 20–21 µm range as compared to the other assays. Length measurements using this method are not optimized for precision, but this method does give a simple way to see the relative size distribution differences in the assays.


Effects of surface passivation on gliding motility assays.

Maloney A, Herskowitz LJ, Koch SJ - PLoS ONE (2011)

Relative microtubule length distributions.Filled blue bars are length calculations for the alpha casein                            passivation, filled green bars are for the beta casein passivation,                            filled red bars are for the kappa casein passivation, unfilled black                            bars are for the mixed casein passivation, and unfilled purple bars are                            for the whole casein passivation. Length measurements were performed                            only on tracked microtubules and are estimations from computing the                            Convex Hull perimeter on eroded images of microtubules.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3108588&req=5

pone-0019522-g002: Relative microtubule length distributions.Filled blue bars are length calculations for the alpha casein passivation, filled green bars are for the beta casein passivation, filled red bars are for the kappa casein passivation, unfilled black bars are for the mixed casein passivation, and unfilled purple bars are for the whole casein passivation. Length measurements were performed only on tracked microtubules and are estimations from computing the Convex Hull perimeter on eroded images of microtubules.
Mentions: Figure 2 shows a histogram of microtubule lengths. To obtain lengths, we used an erosion algorithm on binary images of only the microtubules that were tracked. We used a standard function in LabVIEW/Vision 7.1 called Skeleton L. After the erosion, we used a Convex Hull perimeter calculation also found in the LabVIEW/Vision 7.1 library of functions. We divided the Convex Hull perimeter by 2 to estimate the microtubule length. Two of the authors independently and manually estimated the length of several microtubules and found that usually the Convex Hull perimeter was in between both manually obtained values. We decided that this was sufficient for identifying large changes in MT length distributions, and so we did not further investigate the robustness or systematic errors in our length estimation. As can be seen in the figure, alpha casein (filled blue bars), whole casein (empty purple line bars) and mixed casein (empty black line bars) all were able to sustain motility with smaller microtubules 2–6 µm range. Beta casein (filled green bars) had significantly fewer microtubules that were tracked, however, those that were tended to be longer than the ones tracked in the alpha casein assay. Kappa casein (filled red bars) show that there were many more trackable microtubules than beta casein and significantly fewer than alpha casein. The smallest trackable microtubules using our algorithm are not smaller than 2 µm. Figure 2 shows that while in comparison to the other lengths, there are relatively few 2 µm microtubules in any of the assays, kappa casein had no microtubules that fell in the 2–3 µm range. There is, however, a larger number of kappa casein microtubules that were in the 20–21 µm range as compared to the other assays. Length measurements using this method are not optimized for precision, but this method does give a simple way to see the relative size distribution differences in the assays.

Bottom Line: Beta casein did not support motility very well and averaged speeds of 870±30 nm/s.Kappa casein supported motility very poorly and we were unable to obtain an average speed.Finally, we observed that mixing alpha, beta, and kappa casein with the proportions found in bovine whole casein supported motility and averaged speeds of 966±6 nm/s.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy and Center for High Technology Materials, University of New Mexico, Albuquerque, New Mexico, United States of America. amaloney@unm.edu

ABSTRACT
In this study, we report differences in the observed gliding speed of microtubules dependent on the choice of bovine casein used as a surface passivator. We observed differences in both speed and support of microtubules in each of the assays. Whole casein, comprised of α(s1), α(s2), β, and κ casein, supported motility and averaged speeds of 966±7 nm/s. Alpha casein can be purchased as a combination of α(s1) and α(s2) and supported gliding motility and average speeds of 949±4 nm/s. Beta casein did not support motility very well and averaged speeds of 870±30 nm/s. Kappa casein supported motility very poorly and we were unable to obtain an average speed. Finally, we observed that mixing alpha, beta, and kappa casein with the proportions found in bovine whole casein supported motility and averaged speeds of 966±6 nm/s.

Show MeSH