DEVICES, SYSTEMS AND METHODS FOR BIOMARKER ANALYSIS
Provided herein are devices, systems, kits and methods for predicting or determining the gender of a fetus using cell free fetal nucleic acids in a small amount of maternal biological sample. Devices can be used at point of need during early stages of pregnancy and are compatible with communication devices.
1 ) A device comprising:
a) a sample purifier for removing a cell from a biological fluid sample to produce a cell-depleted sample; and
b) at least one of a detection reagent and a signal detector for detecting a plurality of cell-free DNA fragments in the cell-depleted sample.
2 ) The device of claim 1 , wherein a first sequence is present on a first cell-free DNA fragment of the plurality of cell-free DNA fragments and a second sequence is present on a second cell-free DNA fragment of the plurality of cell-free DNA fragments, and wherein the first sequence is at least 80% identical to the second sequence.
3 ) The device of claim 2 , wherein the device comprises at least one nucleic acid amplification reagent and a single pair of primers capable of amplifying the first sequence and the second sequence.
4 ) The device of claim 2 , wherein at least one of the first sequence and the second sequence is repeated at least twice in a genome of a subject.
5 ) The device of claim 2 , wherein the first sequence and the second sequence are each at least 10 nucleotides in length.
6 ) The device of claim 2 , wherein the first sequence is on a first chromosome and the second sequence is on a second chromosome.
7 ) The device of claim 2 , wherein the first sequence and the second sequence are on the same chromosome but separated by at least 1 nucleotide.
8 ) The device of claim 2 , wherein the first sequence and the second sequence are in functional linkage.
9 ) The device of claim 1 , wherein the sample purifier comprises a filter, and wherein the filter has a pore size of about 0.05 microns to about 2 microns.
10 ) The device of claim 9 , wherein the filter is a vertical filter.
11 ) The device of claim 1 , wherein the sample purifier comprises a binding moiety selected from an antibody, antigen binding antibody fragment, a ligand, a receptor, a peptide, a small molecule, and a combination thereof.
12 ) The device of claim 11 , wherein the binding moiety is capable of binding an extracellular vesicle.
13 ) The device of claim 2 , wherein the at least one nucleic acid amplification reagent comprises an isothermal amplification reagent.
14 ) The device of claim 1 , wherein the signal detector is a lateral flow strip.
15 ) The device of claim 1 , wherein the device is contained in a single housing.
16 ) The device of claim 1 , wherein the device operates at room temperature.
17 ) The device of claim 1 , wherein the device is capable of detecting the plurality of biomarkers in the cell-depleted sample within about five minutes to about twenty minutes of receiving the biological fluid.
18 ) The device of claim 1 , comprising a communication connection.
19 ) The device of claim 1 , comprising a transdermal puncture device.
20 ) A method comprising:
a) obtaining a fluid sample from a subject, wherein the volume of the biological sample is not greater than about 120 microliters;
b) contacting at least one cell free nucleic acid in the fluid sample with an amplification reagent and an oligonucleotide primer that anneals to a sequence corresponding to a sequence of interest in order to produce an amplification product; and
c) detecting the presence or absence of the amplification product, wherein the presence or absence indicates a health status of the subject.
21 ) The method of claim 20 , wherein the fluid sample is a blood sample.
22 ) The method of claim 20 , wherein the fluid sample is a plasma sample from blood.
23 ) The method of claim 22 , wherein the volume of the plasma sample is not greater than 50 μl.
24 ) The method of claim 22 , wherein the volume of the plasma sample is between about 10 μl and about 40 μl.
25 ) The method of claim 20 , wherein the sample contains about 25 pg to about 250 pg of total circulating cell free DNA.
26 ) The method of claim 20 , wherein the sample contains about 5 to about 100 copies of the sequence of interest.
27 ) The method of claim 26 , wherein the copies are at least 90% identical to one another.
28 ) The method of claim 20 , wherein the sequence of interest is at least 10 nucleotides in length.
29 ) The method of claim 20 , wherein contacting comprises performing isothermal amplification.
30 ) The method of claim 20 , wherein contacting occurs at room temperature.
31 ) The method of claim 20 , wherein the method comprises incorporating a tag into the amplification product as the amplifying occurs, and wherein detecting the presence of the amplification product comprises detecting the tag.
32 ) The method of claim 31 , wherein the tag does not comprise a nucleotide.
33 ) The method of claim 31 , wherein detecting the amplification product comprises contacting the amplification product with a binding moiety that is capable of interacting with the tag.
34 ) The method of claim 33 , comprising contacting the amplification product with the binding moiety on a lateral flow device.
35 ) The method of claim 20 , wherein the steps (a) through (c) are performed in less than fifteen minutes.
36 ) The method of claim 20 , wherein the method is performed by the subject.
37 ) The method of claim 20 , wherein the method is performed by an individual without receiving technical training for performing the method.
38 ) The method of claim 20 , wherein obtaining, contacting, and detecting is performed with a single handheld device.
39 ) The method of claim 20 , wherein the health status is selected from the presence and the absence of a pregnancy.
40 ) The method of claim 20 , wherein the health status is selected from the presence and the absence of a neurological disorder, a metabolic disorder, a cancer, an autoimmune disorder, an allergic reaction, and an infection.
41 ) The method of claim 20 , wherein the health status is a response to a drug or a therapy.
42 ) A device comprising:
a) a sample purifier that removes a cell from a fluid sample of a female subject;
b) at least one nucleic acid amplification reagent;
c) at least one oligonucleotide comprising a sequence corresponding to a Y chromosome, wherein the at least one oligonucleotide and nucleic acid amplification reagent are capable of producing an amplification product; and
d) at least one of a detection reagent or a signal detector for detecting the amplification product.
43 ) The device of claim 11 , wherein the oligonucleotide comprises a sequence corresponding to a gene selected from DYS14 gene or a TTTY22.
44 ) A method comprising:
a) obtaining a fluid sample from a female pregnant subject, wherein the volume of the biological sample is not greater than about 300 microliters;
b) contacting at least one cell free nucleic acid in the fluid sample with an amplification reagent and an oligonucleotide primer that anneals to a sequence corresponding to a sex chromosome; and
c) detecting the presence or absence of an amplification product, wherein the presence or absence indicates the gender of a fetus of the female pregnant subject.
45 ) The method of claim 43 , wherein the fluid sample is a blood sample.
46 ) The method of claim 44 , wherein the volume of the blood sample is not greater than 120 μl.
47 ) The method of claim 43 , wherein the fluid sample is a plasma sample from blood.
48 ) The method of claim 46 , wherein the volume of the plasma sample is not greater than 50 μl.
49 ) The method of claim 46 , wherein the volume of the plasma sample is between about 10 μl and about 40 μl.
50 ) The method of any one of claims 43 to 48 , wherein obtaining comprises performing a finger prick.