In-vitro contractile force indicator
Devices and methods to measure cells and/or tissue's contractile force are disclosed. Included is a mount with rigid, and non-rigid posts sized to flex. Determined is force exerted by contractile cells and tissues in a multiwell plate. The device is designed to fit inside individual wells with posts directed downwards. Posts are attached to a 3D printed circular mount with tabs for depth within the well. The mount has a window for medium changes while the device is positioned inside the well. The cells are seeded within a hydrogel. As the hydrogel condenses, cells/tissue wrap around the post's outside pulling non-rigid post toward rigid post. Inverted light microscope is used to determine deflection of non-rigid post inside the multiwell plate. Movement of the non-rigid post is measured using an acrylic ruler on an underside of the multiwell plate. Contractile forces of cells/tissue are determined using cantilever mechanics.
1. A contractile force indicator comprising:
a first mount having a cylindrical shape to fit inside a well of a culture plate that contains a growing specimen;
a first rigid post defined as having no deflection or no deformation when an external force of at least 30 μN is applied by the growing specimen, the first rigid post is integral to the first mount and the first rigid post remains stationary with respect to the first mount; the first rigid post is in a vertical position relative to the first mount; and the first rigid post is pointing in a downward direction into the well;
a first non-rigid post defined as having a deflection or a deformation when the external force of at least 30 μN is applied by the growing specimen; the first non-rigid post is interchangeably attached to the first mount and parallel to the first rigid post; and the first non-rigid post is pointing in the downward direction into the well;
wherein the first rigid post serves as an anchor when the first non-rigid post undergoes a deflection due to the external force of at least 30 μN applied by the specimen contracting and wrapping around the first rigid and the first non-rigid posts; and
wherein the deflection is for calculating a force of contraction of the specimen.
2. The contractile force indicator in claim 1 , further comprising:
a ruler for measuring the deflection;
wherein the culture plate is a multiwell culture plate and the ruler is disposed underneath the multiwell culture plate during measurement;
the ruler further having a plurality of circular patterns to fit under each well of the multiwell culture plate; and
the plurality of circular patterns each further having a parallel pattern of hatch marks for measuring the deflection of the first non-rigid post.
3. The contractile force indicator of claim 1 , further comprising:
a second mount having a second rigid post; the second mount connected to the first mount by a bridge portion;
wherein the first and the second mounts, the first and the second rigid posts, and the bridge portion are all integral; and
wherein the first and second mounts are inside two adjacent wells within a multiwell culture plate to prevent rotation or movement during culture of the specimen.
4. The contractile force indicator of claim 3 , wherein the second mount includes a second non-rigid post that is a different dimension or a different material than the first non-rigid post.
5. The contractile force indicator of claim 3 , wherein the second mount includes a second non-rigid post that is the same dimension as the first non-rigid post.
6. The contractile force indicator of claim 3 , wherein the first mount further includes a first support and a second support; the first support having the first rigid post integrally attached thereon; the second support having a hole for interchangeable or for fixable attachment of the first non-rigid post.
7. The contractile force indicator of claim 6 , wherein the first and the second supports define a window;
the window for allowing medium changes in each well of the multiwell culture plate without removal of the first mount from the multiwell culture plate; and the window for observing a ruler underneath the multiwell culture plate wherein the ruler has a circular pattern under each well; and the circular pattern further has a parallel pattern of hatch marks or vertical lines for measuring the deflection; and
wherein the vertical lines of the ruler having a width of 0.035 mm and spaced 1.00 mm apart.
8. The contractile force indicator of claim 7 , wherein the ruler is transparent and allows bottom up imaging of the specimen in each well, wherein imaging is done starting at the bottom of each well in the multiwell culture plate.
9. The contractile force indicator of claim 8 , wherein the specimen is a plurality of cells or tissue in a medium, wherein the cells or tissue contains myoblasts isolated from patients presenting with Duchenne's muscular dystrophy, and wherein the plurality of cells exert contractile forces, and the contractile force indicator measures effects of different drugs or drug dosages on muscular strength or recovery.
10. The contractile force indicator of claim 3 , wherein each well of the multiwell culture plate contains varying hydrogel compositions; or varying cellular composition or phenotypes.
11. The contractile force indicator of claim 1 , wherein the first mount, and the first rigid post are made of a mixture of 2-hydroxyethyl methacrylate, phenyl bis(2,4,6-trimethyl benzoyl)-phosphine oxide, and urethane dimethacrylate and the first non-rigid post is made of polydimethylsiloxane (PDMS) that are both autoclavable.
12. The contractile force indicator of claim 11 , wherein the PDMS has an elastic modulus of 1.20±0.60 MPa after 1× autoclave.
13. The contractile force indicator of claim 1 , wherein the specimen contains cells; and the deflection caused by cell forces constricting the first rigid and the first non-rigid posts as the cells grow is measured in real time.
14. The contractile force indicator of claim 1 , where the first mount further includes a plurality of tabs to allow the first mount to sit in a multiwell culture plate; and dispose the first rigid and the first non-rigid posts into the specimen at the same distance.
15. The contractile force indicator of claim 1 , wherein the first rigid post is 10 mm long with a diameter of 2 mm, and the first non-rigid post is 10 mm long.
16. The contractile force indicator of claim 15 , wherein the diameter of the non-rigid post is changeable to match a strength of the specimen and wherein the specimen is contractile tissue.
17. The contractile force indicator of claim 16 , wherein the non-rigid post is 1 mm diameter.
18. The contractile force indicator of claim 1 , wherein the first mount and the first rigid post are 3D printed, and the first non-rigid post is cast in a mold.