Team:HKU-HKBU/Motor Preliminary Trials

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Revision as of 15:13, 19 October 2009

Membrane Version

We used Immobilon-P transfer membrane as the material for motor. Before cutting down the membrane into small pieces, pre-activation should be applied to it first [Link 1]. After the pre-activation, the membrane was first sheared into strings manually by using scissors, with the width and thickness both around 100μm. Then, we used Leica-crytomicrotome to cut the “threads” into small fragments, with the length of 60μm. Thus, we got the motors, whose demention was aproximately 100μm×60μm×100μm. Since the surface of membrane had already been activated, the polar-expression bacteria could bind onto such biotin-coated motors.

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 {{Team:HKU-HKBU/header}}
-==Membrane Production==
+==Membrane Version==
-===Step 1===
+We used Immobilon-P transfer membrane as the material for motor. Before cutting down the membrane into small pieces, pre-activation should be applied to it first [Link 1]. After the pre-activation, the membrane was first sheared into strings manually by using scissors, with the width and thickness both around 100μm. Then, we used Leica-crytomicrotome to cut the “threads” into small fragments, with the length of 60μm. Thus, we got the motors, whose demention was aproximately 100μm×60μm×100μm. Since the surface of membrane had already been activated, the polar-expression bacteria could bind onto such biotin-coated motors.
-The Immobolin-P membrane was first made wet and consequently homogenized with a mini-homogenizer.
-Results: The membrane could not be broken into small pieces.
-[[Image:HKU-HKBU_motor_results_1.png | none |thumb]]
-===Step 2===
-The Immobolin-P membrane was first moistened, then it was put into a 10ml centrifugation tube. The tube was then totally filled with glass beads, and undergo vortexing subsequently.
-Results: The membrane remained intact.
-[[Image:HKU-HKBU_motor_results_2.png | none|thumb]]
-[[Image:HKU-HKBU_motor_results_3.png | none |thumb]]
-===Step 3===
-The Immobolin-P membrane was moistened and liquefied nitrogen was poured onto it. The membrane was broken into pieces by human hands.
-Results: The membrane remained intact.
-===Step 4===
-The Immobolin-P membrane was first biotinylated and cut into very small pieces by human hands. Then, the membrane fragments were put into a mould made with aluminium foil and fixed into it with the help of glue. The mould along with the membrane fragment were cut with a Leica-crytomicrotome into further smaller pieces the size of 100umx60umx100um.
-[[Image:HKU-HKBU_motor_results_4.png | none|thumb]]
-[[Image:HKU-HKBU_motor_results_5.png | none |thumb]]
-[[Image:HKU-HKBU_motor_results_6.png | none|thumb]]
-===Step 5===
-Some elemental silicon fragments were put inside a 1-ml eppendorf tube. The protein-biotin complex and some concentrated HCL were then added into the same tube. The tube was put inside a water bath for 2 hours. The silicon fragments were made dry by rinsing with PBS and followed by air drying. Then, some streptavidin containing beads were added onto the fragments. The fragments were observed under a microscope.
-Results: No beads were bound to the silicon fragments.
-===Step 6===
-The silicon fragments were silanized by soaking them for 2 h in a solution of aminopropyl triethoxyl silane (3% aminopropyl triethoxyl silane, 2% acetic acid, 5% water, 90% ethanol), then rinsed with ethanol, and dried with a PCR machine for 5 mins. The amino-coated rotors (Fig. 2Bh) were then reacted with 1 mM succinimidyl-6-(biotinamido)-6-hexana- mido hexanoate (EZ-Link NHS-LC-LC-biotin; Pierce, Rock- ford, IL) dissolved in 40 mM phosphate buffer (pH 8.0) for 1 h at 37°C. Then strepatavidin beads were allowed to bind onto it and the fragments were observed under a microscope.
 {{Team:HKU-HKBU/footer}}