CONTRACTILITY ASSAY
Cells are magnetized and then grown in ring shaped 3D culture using a ring magnet. Contractility is measuring by tracking the size of the hole in the 3D cellular ring.
1 ) A contractility assay comprising:
a) obtaining contractile cells;
b) magnetizing said contractile cells;
c) magnetically creating a 3D ring of magnetized cells using a ring magnet, said 3D ring having a hole;
d) taking photomicrographs of said 3D ring of magnetized cells at least one or more times; and,
e) analyzing said photomicrographs to measure hole size or rate of change of hole size or both;
f) wherein hole size hole size or rate of change of hole size relates to contractility of said contractile cells.
2 ) The method of claim 1 , further including preparing a plurality of 3D rings for use as control samples and a preparing a plurality of 3D rings for use as test samples, wherein a test agent is added to each of said test samples.
3 ) The method of claim 2 , further including adding varying amounts of said test agent to said plurality of test samples.
4 ) The method of claim 3 , comprising taking a plurality of photomicrographs of said plurality of test samples and said control samples at a plurality of times.
5 ) The method of claim 3 , further comprising washing out said test agent and taking a further plurality of photomicrographs of said test samples and said control samples at a further plurality of times.
6 ) The method of claim 1 , wherein cells are magnetized with a composition comprising: a) a negatively charged nanoparticle; b) a positively charged nanoparticle; and c) a support molecule, wherein one of said negatively charged nanoparticle or positively charged nanoparticle contains a magnetically responsive element, and wherein said support molecule holds said negatively charged nanoparticle and said positively charged nanoparticle in an intimate and disordered admixture, not a micelle.
7 ) The method of claim 6 , wherein the support molecule comprises peptides, polysaccharides, nucleic acids, polymers, poly-lysine, fibronectin, collagen, laminin, BSA, hyaluronan, glycosaminoglycan, anionic, non-sulfated glycosaminoglycan, gelatin, nucleic acid, extracellular matrix protein mixtures, antibody, or mixtures or derivatives thereof, wherein said negatively charged nanoparticle is a gold nanoparticle, and wherein said positively charged nanoparticle is an iron oxide nanoparticle.
8 ) The method of claim 6 , wherein the composition comprises poly-lysine, gold nanoparticles, and iron oxide nanoparticles.
9 ) The method of claim 1 , wherein said contractile cells are obtained from a patient.
10 ) The method of claim 1 , wherein said contractile cells are obtained from a maternal patient's uterus, myometrium, placenta, vasculature, or umbilical cord.
11 ) The method of claim 1 , wherein said contractile cells are obtained from a patient's uterus.
12 ) A uterine contractility assay comprising:
a) obtaining smooth muscle cells (SMCs) from a uterus, myometrium, placenta, maternal vasculature, or umbilical cord;
b) magnetizing said SMCs to make magnetized cells;
c) magnetically creating a 3D ring of magnetized cells using a ring magnet, said 3D ring having a hole;
d) adding a test agent to said 3D ring of magnetized cells;
e) taking photographs of said 3D ring of magnetized cells at one or more times before and after adding said test agent; and,
f) analyzing said photographs to measure rate of change of hole size;
g) wherein rate of change of hole size relates to contractility of said SMCs in response to said test agent.
13 ) The method of claim 12 , further including preparing a plurality of 3D rings for use as control samples and preparing a plurality of 3D rings for use as test samples, wherein a test agent is added to each of said test samples.
14 ) The method of claim 13 , further including adding varying amounts of said test agent to said plurality of test samples.
15 ) The method of claim 14 , comprising taking a plurality of photographs of said 3D rings at a plurality of times.
16 ) The method of claim 12 , further comprising washing out said test agent and taking a further plurality of photographs of said 3D rings at a further plurality of times.
17 ) The method of claim 12 , wherein said SMCs are magnetized with a composition comprising: a) a negatively charged nanoparticle; b) a positively charged nanoparticle; and c) a support molecule, wherein one of said negatively charged nanoparticle or positively charged nanoparticle contains a magnetically responsive element, and wherein said support molecule holds said negatively charged nanoparticle and said positively charged nanoparticle in an intimate admixture, not a micelle.
18 ) The method of claim 17 , wherein the composition comprises poly-lysine, gold nanoparticles, and iron oxide nanoparticles.
19 ) The method of claim 11 , wherein contraction is initiated by removing said ring magnet.
20 ) The method of claim 11 , wherein contraction is initiated by adding a contractile agent or combination of agents.
21 ) An assay device comprising a microtiter plate having a plurality of wells, each well containing a culture medium containing a 3D ring of magnetic contractile cells.
22 ) The device of claim 21 , wherein each said 3D ring of magnetic contractile cells is floating in said culture medium.
23 ) The device of claim 21 , wherein said magnetic contractile cells are myometrial smooth muscle cells.
24 ) A uterine contractility assay comprising:
a) obtaining contractile cells from a myometrium of a patient's uterus, and optionally freezing said cells before step b);
b) allocating said contractile cells to a plurality of wells in a microtiter plate;
c) magnetizing said contractile cells;
d) magnetically creating a 3D ring of magnetized cells using a ring magnet, said 3D ring having a hole, said hole having a size;
e) adding a test agent to said 3D ring of magnetized cells;
f) initiating contraction by removing said ring magnet; either before or after step e);
g) taking photographs of an entirety of said microtiter plate at one or more times before and after adding said test agent; and,
h) analyzing said photographs to measure a rate of hole size contraction;
i) wherein a reduced rate of hole size contraction means said test agent inhibits contraction.
25 ) The method of claim 24 , wherein said microtiter plate is a 384-well microtiter plate, and wherein 10-100×10 3 cells/well are used to form said 3D ring.
26 ) The method of claim 24 , wherein said cells are grown in 3D culture for 1-7 days to form a 3D spheroid and wherein said spheroid is broken up to provide contractile cells for said allocating step b.
27 ) The method of claim 24 , wherein said 3D rings are cultured for 1-3 days before said adding step e.