High-Throughput Yeast-Aging Analysis (HYAA) Chip For Performing Yeast Aging Assays

Patent metadata

Field	Value
Publication number	US-2016281126-A1
Application number	US-201615173713-A
Country	US
Kind code	A1
Filing date	Jun 5, 2016
Priority date	Jun 5, 2016
Publication date	Sep 29, 2016
Grant date	—

Abstract

An improved technique for studying the molecular mechanisms of aging in eukaryotic cells utilizes an efficient, high-throughput microfluidic single-cell analysis chip in combination with high-resolution time-lapse microscopy. A High-throughput Yeast Aging and Analysis (HYAA) Chip has a plurality of discrete microfluidic channels grouped into a number of modules. Each module has a single medium inlet and a single medium outlet. Each channel in a module has a microfluidic chamber having a plurality of single-cell trapping structures, and features a sample inlet for introducing cells into the flow of medium through the chamber. This innovative design enables the determination of the yeast replicative lifespan in a high throughput manner.

First claim

What is claimed is:

1 . A module for isolating and culturing a plurality of single cells, comprising:
a thin sheet of flexible or semi-rigid material; a medium inlet, a medium outlet, and a channel extending in fluid communication between the medium inlet and the medium outlet; a chamber disposed the channel; a plurality of single-cell trapping structures disposed in the chamber; and a sample inlet for introducing cells into a flow of medium through the chamber.

2 . The module of claim 1 , wherein the thin sheet of flexible or semi-rigid material is selected from the group consisting of polydimethylsiloxane (PDMS), PMMA (poly(methyl methacrylate)), PS (polystyrene), and PC (polycarbonate).

3 . The module of claim 2 , wherein the thin sheet of flexible or semi-rigid material has a thickness of approximately 8 μm.

4 . The module of claim 2 , wherein the thin sheet of flexible or semi-rigid material is molded to have channels and single-cell trapping structures on a front surface thereof.

5 . A high-throughput yeast-aging analysis (HYAA) chip comprising a plurality of modules according to claim 1 , wherein:
each module has a single medium inlet disposed on one side thereof, and a single medium outlet disposed on an opposite side thereof.

6 . The HYAA chip of claim 5 , wherein:
the modules are disposed one above the other on a surface of the thin sheet of flexible or semi-rigid material so that their inlets are all oriented to the one side of the sheet and their outlets are oriented to an opposite side of the sheet.

7 . The HYAA chip of claim 5 , wherein:
each module includes a plurality of channels branched from its single medium inlet and merged into its single medium outlet.

8 . The HYAA chip of claim 7 , wherein:
the channels are parallel with one another.

9 . The HYAA chip of claim 5 , further comprising:
a plurality of single-cell trapping structures disposed in each of the chambers, so as to be capture single cells from the flow of medium through the chamber.

10 . The HYAA chip of claim 9 , wherein:
the trapping structures are arranged in an array having a number of columns of trapping structures disposed vertically, one above the other, with spaces therebetween.

11 . The HYAA chip of claim 10 , wherein:
the trapping structures of one column are offset vertically from the trapping structures of an adjacent column to facilitate flow of medium and cells through the array.

12 . The module of claim 1 , wherein:
the trapping structures are cup-shaped, having an inlet and an outlet; wherein the inlets of the trapping structures are larger than the outlets of the trapping structures.

13 . The module of claim 12 , wherein:
a width of the inlets of the trapping structures is approximately 6 μm; a width of the outlets of the trapping structures is approximately 3 μm.

14 . The module of claim 13 , wherein:
a height of the trapping structures is approximately 5 μm.

15 . The module of claim 14 , wherein:
a plurality of trapping structures are arranged in an array of columns and rows; and a column spacing is equal to or smaller than a row spacing to ensure high trapping efficiency and minimal channel obstruction by daughter cells removed from the trapped mother cells.

16 . The module of claim 1 , wherein:
the dimensions of the trapping structures are optimized to ensure that at least one of the following conditions are met: (i) only a single cell is captured in each trapping structure; (ii) the trapped cells are stably retained in the trapping structure during the entire course of an aging experiment; and (iii) the trapping structure does not pose a spatial constraint to cell size increase during aging.

17 . A method of isolating and culturing a plurality of single cells, comprising:
providing a module comprising:
a medium inlet, a medium outlet, and a channel extending in fluid communication between the medium inlet and the medium outlet;
a chamber disposed the channel;
a plurality of single-cell trapping structures disposed in the chamber; and
a sample inlet for introducing cells into a flow of medium through the chamber; and
further comprising: introducing a liquid medium continuously through the medium inlet; injecting suspended yeast cells through the sample inlet; and trapping individual mother cells in the single-cell trapping structures.

18 . The method of claim 17 , further comprising:
cultivating the trapped mother cells with continuous medium flow.

19 . The method of claim 18 , further comprising:
as the trapped cells mother cells grow and bud, and daughter cells are produced and detached from their mother cells, removing the daughter cells by the medium flow.

20 . The method of claim 19 , further comprising:
tracking the development of the mother cells over their entire lifespan in a single experiment using high-resolution multi-position time-lapse microscopy.

Assignees

• Qin Lidong
• Baylor College Medicine

Inventors

• Qin Lidong
• Chan Jo Myeong
• Dang Weiwei

Classifications

• B01L2300/0864

  comprising only one inlet and multiple receiving wells, e.g. for separation, splitting · CPC title

• B01L2300/0819

  Microarrays; Biochips · CPC title

• B01L3/502761

  specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads or physically stretching molecules · CPC title

• B01L2300/0867

  Multiple inlets and one sample wells, e.g. mixing, dilution · CPC title

• B01L3/502715

  characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces · CPC title