DIRECT IONIZATION IN IMAGING MASS SPECTROMETRY OPERATION
As described herein, one or more parameters of a direct ionization imaging mass spectrometer (IMS) may be set to obtain a desired plasma and deliver it to a mass detector. Depending on the application, certain parameters may be predetermined (e.g., a spot size given a desired resolution) and, as described herein, other parameters can be adjusted to obtain the desired plasma properties. Also included is sample preparation suitable for direct ionization IMS and/or other imaging modalities.
1 . A method of analyzing a sample comprising:
d) directing radiation at a spot on a sample to form a plasma comprising elemental ions,
e) delivering the elemental ions to a mass detector;
f) detecting the elemental ions at the mass detector.
2 . The method of claim 1 , further comprising an initial step of providing a sample on a solid support.
3 . The method of claim 1 , wherein the sample is a geological or semiconductor sample.
4 . The method of claim 1 , wherein sample is a biological sample.
5 . The method of claim 4 , wherein the sample is a tissue section, such as an EM section.
6 . The method of claim 5 , wherein the tissue section is 100 nm thick or less.
7 . The method of claim 4 , wherein the sample is stained with specific binding partners (SBPs) comprising distinct metal tags.
8 . The method of claim 7 , wherein the SBPs are antibodies.
9 . The method of claim 7 , further comprises metal containing histochemical stains and/or metal tagged oligonucleotides.
10 . The method of claim 2 , wherein the solid support comprises an X-Y stage.
11 . The method of claim 2 , wherein the solid support comprises a slide.
12 . The method of claim 1 , wherein the radiation is scanned across the sample.
13 . The method of claim 12 , wherein the radiation is laser radiation scanned across the sample by a positioner.
14 . The method of claim 13 , wherein the positioner is a galvanometer mirror, piezoelectric mirror, MEMS mirror, polygon scanner, acousto-optic device or an electro-optic device.
15 . The method of claim 1 , wherein the radiation is directed from a different angle than the direction of the mass detector in the relation to the sample.
16 . The method of claim 15 , wherein the radiation is directed from the opposite side of the sample from the side of delivery to the mass detector.
17 . The method of claim 1 , wherein the radiation is a laser.
18 . The method of claim 17 , wherein the laser has a pulse duration between 10 fs and 10 ps.
19 . The method of claim 17 , wherein the laser has a pulse duration less than 10 ps.
20 . The method of claim 17 , wherein the laser is a high harmonic generation laser.
21 . The method of claim 17 , wherein the laser is focused by an immersion lens.
22 . The method of claim 21 , wherein the laser is focused by a liquid or solid immersion lens.
23 . The method of claim 17 , wherein the laser is from a femtosecond laser or picosecond laser.
24 . The method of claim 17 , wherein the laser has a wavelength of less than 500 nm.
25 . The method of claim 17 , wherein the laser is a UV laser or EUV laser.
26 . The method of claim 17 or 18 , wherein the laser has a pulse energy between 10 pj and 10 uJ.
27 . The method of claim 17 , wherein the laser has a pulse energy of less than 1 nj.
28 . The method of claim 27 , wherein the laser has a pulse energy of less than 100 pj.
29 . The method of claim 1 , wherein the radiation is a beam of charged particles.
30 . The method of claim 29 , wherein the charged particle beam is an electron beam.
31 . The method of claim 30 , wherein the electron beam comprises electrons with an energy of between 100 eV, and 10 keV.
32 . The method of claim 30 or 31 , wherein the number of electrons used to create the plasma is at or between 1000 and 50000 electrons
33 . The method of claim 1 , wherein the radiation is a pulse of radiation of a duration less than the time of plasma formation
34 . The method of claim 1 , wherein sample spots are analyzed at a frequency between 1 kHz and 10 MHz.
35 . The method of claim 1 , 18 or 26 , wherein the spot size is 300 nm or less.
36 . The method of claim 35 , wherein the spot size is 100 nm or less.
37 . The method of claim 36 , wherein the spot size is 50 nm or less.
38 . The method of claim 37 , wherein the spot size is 30 nm or less.
39 . The method of claim 1 , wherein the ions are delivered in a vacuum from the point of plasma formation.
40 . The method of claim 1 , wherein the plasma is not formed in the presence of an injected noble gas, such as Argon or Xenon.
41 . The method of claim 1 , wherein the plasma is a thermal plasma, having an internal temperature between 3000 and 30000 K.
42 . The method of claim 41 , wherein the internal temperature is between 5000 and 10000 K.
43 . The method of claim 41 , wherein the thermal plasma internal temperature is within 3000 to 30000 K past neutralization.
44 . The method of claim 1 , wherein the plasma is a non-thermal plasma.
45 . The method of claim 1 , wherein the plasma has a diameter less than 1 um when it passes the point of neutralization.
46 . The method of claim 1 , wherein the elemental ions from plasma are directly delivered to the mass detector by ion transport optics.
47 . The method of claim 1 , wherein delivering does not comprise a mass filter.
48 . The method of claim 46 , wherein the ion transport optics comprises a high pass filter with a cutoff below 80 amu.
49 . The method of claim 1 , wherein the delivery time of elemental ions from the plasma to the detector is less than 200 us.
50 . The method of claim 1 , wherein at least 10% of metals released from the sample spot by the radiation are atomized and ionized and delivered to the detector.
51 . The method of claim 1 , wherein the ionization efficiency of lanthanides is in the plasma is at least 20% and the ionization efficiency of carbon in the plasma is below 5%.
52 . The method of claim 1 , wherein the plasma has an ionization efficiency of at least 5% post neutralization.
53 . The method of claim 1 , wherein the detector is a magnetic sector detector.
54 . The method of claim 1 , wherein the detector is a TOF detector.
55 . The method of claim 54 , wherein ions from a single spot are not separately pushed to the TOF detector.
56 . The method of claim 1 , wherein detection of the elemental ions comprises analysis of metal tags or targets associated with the metal tags.
57 . The method of claim 1 , further comprising forming an image of the sample based on the elemental/isotopic composition of multiple spots.
58 . The method of claim 4 , further comprising detecting single copies of metal-tagged antibodies.
59 . The method of claim 58 , wherein at least some of the metal-tags comprise a barcode of isotopes.
60 . The method of claim 59 , wherein the sample comprises more than 100 different metal tagged antibodies
61 . The method of claim 1 , wherein the portion of the sample removed at the spot by radiation is less than 1 atto gram.
62 . The method of claim 1 , further comprising 3D imaging by radiating the sample at the same X, Y coordinate multiple times.
63 . A system for analyzing a sample comprising:
a) a solid support;
b) a radiation source and optics for directing radiation at a spot on a sample to form a plasma that atomizes and ionizes the sample at that spot to produce elemental ions;
c) a mass detector for detecting the elemental composition of elemental ions delivered from the plasma.
64 . The system of claim 63 , further comprising a sample mounted on the sample support.
65 . The system of claim 64 , wherein the sample is a geological or semiconductor sample.
66 . The system of claim 64 , wherein sample is a biological sample.
67 . The system of claim 66 , wherein the sample is a tissue section, such as an EM section.
68 . The system of claim 67 , wherein the tissue section is 100 nm thick or less.
69 . The system of claim 66 , wherein the sample is stained with specific binding partners (SBPs) comprising distinct metal tags.
70 . The system of claim 69 , wherein the SBPs are antibodies.
71 . The system of claim 70 , further comprises metal containing histochemical stains and/or metal tagged oligonucleotides.
72 . The system of claim 63 , wherein the solid support comprises an X-Y stage.
73 . The system of claim 63 , wherein the solid support comprises a slide.
74 . The system of claim 63 , wherein the radiation source is a laser scanned across the sample by a positioner.
75 . The system of claim 74 , wherein the positioner is a galvanometer mirror, piezoelectric mirror, or MEMS mirror, polygon scanner, acousto-optic device or an electro-optic device.
76 . The system of claim 63 , wherein the radiation source is positioned to direct radiation from a different angle than the direction of the mass detector in the relation to the sample.
77 . The system of claim 76 , wherein the radiation source is positioned to direct radiation from the opposite side of the sample from the side of delivery to the MS detector.
78 . The system of claim 63 , wherein the radiation source is a laser.
79 . The system of claim 78 , wherein the laser has a pulse duration between 10 fs and 10 ps.
80 . The system of claim 78 , wherein the laser has a pulse duration less than 10 ps.
81 . The system of claim 78 , wherein the laser is a high harmonic generation laser.
82 . The system of claim 78 , wherein the laser is focused by an immersion lens.
83 . The system of claim 78 , wherein the laser is focused by a solid or liquid immersion lens.
84 . The system of claim 78 , wherein the laser is from a femtosecond laser or picosecond laser.
85 . The system of claim 78 , wherein the laser has a wavelength of less than 500 nm.
86 . The system of claim 78 , wherein the laser is a UV laser or EUV laser.
87 . The system of claim 78 or 79 , wherein the laser has a pulse energy between 10 pj and 10 uJ.
88 . The system of claim 78 , wherein the laser has a pulse energy of less than 1 nj.
89 . The system of claim 88 , wherein the laser has a pulse energy of less than 100 pj.
90 . The system of claim 63 , wherein the radiation is a beam of charged particles.
91 . The system of claim 90 , wherein the beam of charged particles is an electron beam.
92 . The system of claim 91 , wherein the electron beam can direct electrons with an energy of between 100 eV, and 100 keV to the sample spot.
93 . The system of claim 30 or 31 , wherein the radiation source can direct a number of electrons to the sample spot at or between 1000 and 50000 electrons.
94 . The system of claim 63 , wherein the radiation source is configured to provide a pulse of radiation of a duration less than the time of plasma formation.
95 . The system of claim 63 , wherein sample spots are analyzed at a frequency between 1 kHz and 10 MHz.
96 . The system of claim 63 , 78 , or 88 , wherein the system comprises radiation optics providing a spot size of 500 nm or less.
97 . The system of claim 96 , wherein the spot size is 200 nm or less.
98 . The system of claim 97 , wherein the spot size is 100 nm or less.
99 . The system of claim 98 , wherein the spot size is 50 nm or less.
100 . The system of claim 63 , wherein the system is configured to maintain a vacuum at the point of plasma formation.
101 . The system of claim 63 , wherein system is configured to form a plasma without the presence of an injected noble gas, such as Argon or Xenon.
102 . The system of claim 63 , wherein the system is configured to form a plasma is a thermal plasma, having an internal temperature between 3000 and 30000K.
103 . The system of claim 102 , wherein the internal temperature is between 5000 and 10000K.
104 . The system of claim 102 , wherein the thermal plasma internal temperature is between 3000 and 30000K past neutralization.
105 . The system of claim 63 , wherein the plasma is a non-thermal plasma.
106 . The system of claim 63 , wherein the plasma has a diameter less than 1 um when it passes the point of neutralization.
107 . The system of claim 63 , wherein the elemental ions from plasma are directly delivered to the mass detector by ion transport optics.
108 . The system of claim 63 , wherein delivering does not comprise a mass filter.
109 . The system of claim 46 , wherein the ion transport optics comprises a high pass filter with a cutoff below 80 amu.
110 . The system of claim 63 , wherein the delivery time of elemental ions from the plasma to the detector is less than 200 us.
111 . The system of claim 63 , wherein at least 10% of metals released from the sample spot by the radiation are atomized and ionized and delivered to the detector.
112 . The system of claim 63 , wherein the ionization efficiency of lanthanides is in the plasma is at least 20% and the ionization efficiency of carbon in the plasma is below 5%.
113 . The system of claim 63 , wherein the plasma has an ionization efficiency of at least 5% post neutralization.
114 . The system of claim 63 , wherein the detector is a magnetic sector detector.
115 . The system of claim 63 , wherein the detector is a TOF detector.
116 . The system of claim 115 , wherein ions from a single spot are not separately pushed to the TOF detector.
117 . The system of claim 63 , wherein detection of the elemental ions comprises analysis of metal tags or targets associated with the metal tags.
118 . The system of claim 63 , wherein the system is configured to form an image of the sample based on the elemental/isotopic composition of multiple spots.