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The nucleus of the cell is a membrane-bound organelle that includes the nuclear envelope, nucleolus, and nuclear matrix, and is the site of gene expression. The nucleus can be selectively visualized by staining nuclear proteins or directly staining nucleic acids. Here we describe a wide selection of nuclear stains that are available with a choice of wavelengths for multiplexing and colocalization in either live or fixed cells.
Cell-permeant nucleic acid stains make it possible to stain live cells or tissues with minimal processing. They reveal the natural location of cells in tissues and provide a means to follow nuclear changes throughout cellular processes, from mitosis to apoptosis. Cell-impermeant nucleic acid stains, used with fixed cells or tissues, are also used as dead-cell indicators providing a means to follow cellular processes, from apoptosis to viability.
The nucleus (plural: nuclei) is found in eukaryotic cells. While most cells have a single nucleus, some cell types do not have a nucleus while others have many nuclei. A cell’s nucleus is a membrane-bound organelle that consists of the nuclear envelope, a double membrane that surrounds and isolates the nuclear contents, and the nuclear matrix which acts like the cytoskeleton and provides support [1,2].
The nucleus contains the cell’s genetic material (chromosomes) and is the primary site of gene expression and DNA replication during cell cycle. The nucleus also contains various proteins such as histones, which form chromosomes.
Within the nucleus is a sub-compartment known as the nucleolus, which is responsible for synthesizing ribosomal RNA (rRNA) and subsequently assembling ribosomes.
ReadyProbes reagents provide the easiest fluorescent staining method for nucleic acids in live or fixed cells. These are ready-to-use solutions in convenient dropper bottles, formulated for room temperature storage.
Learn more about ReadyProbes reagents
Readout |
Fluorescent staining of nucleic acids
| ||
Target |
Membrane-permeable dyes targeting RNA and DNA
|
Membrane impermeable dye targeting RNA and DNA
| |
Common filter set | DAPI | Cy5 | DAPI |
Labels | Hoechst 33342 | NucRed Live | DAPI |
Ex/Em (nm) | 360/460 | 638/686 | 360/460 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
No
|
Fixed cells |
No
|
No
|
Yes
|
Fixable |
Yes
|
No
|
No
|
Platform |
Imaging
|
Imaging
|
Imaging
|
Protocol | Microscopy protocol | Microscopy protocol | Microscopy protocol |
Format |
6 x dropper bottles
|
6 x dropper bottles
|
6 x dropper bottles
|
Cat. No. |
SYTO dyes are live cell, cell-permeant nucleic acid stains that exhibit bright fluorescence upon binding to nucleic acids. Hoechst dyes are blue-fluorescent nucleic acid stains that have multiple applications including distinguishing condensed pycnotic nuclei in apoptotic cells and for cell-cycle studies in combination with BrdU.
Learn more about SYTO nucleic acid stains
Hoechst 33342 | SYTO 9 Green Nucleic Acid Stain | SYTO 82 Orange Nucleic Acid Stain | SYTO 59 Red Nucleic Acid Stain | SYTO Deep Red Nucleic Acid Stain | |
---|---|---|---|---|---|
Readout | Fluorescent staining of nucleic acids | ||||
Target | Preferentially binds AT regions of dsDNA | Cell-permeant dyes targeting DNA and RNA | |||
Common filter set | UV / DAPI | FITC | TRITC | Texas Red | Cy5 |
Labels | Hoechst 33342 | SYTO 9 | SYTO 82 | SYTO 59 | SYTO Deep Red |
Ex/Em (nm) | 350/461 | 483/503 | 541/560 | 622/645 | 652/669 |
Signal-to-noise ratio | |||||
Photostability | |||||
Multiplexing | Yes | Yes | Yes | Yes | Yes |
Live cells | Yes | Yes | Yes | Yes | Yes |
Fixed cells | No | No | No | Yes | No |
Fixable | Yes | Yes | Yes | Yes | Yes |
Platform | Imaging | Imaging | Imaging | Imaging | Imaging |
Protocol | Imaging protocol | — | Imaging protocol | Imaging protocol | — |
Format | 10 mL | 100 μL | 250 µL | 100 µL | 1 vial |
Cat. No. | H3570 | S34854 | S11363 | S11341 | S34900 |
SYTO RNASelect Green Fluorescent Cell Stain is a cell-permeant nucleic acid stain that is selective for RNA. Although virtually nonfluorescent in the absence of nucleic acids, the SYTO RNASelect stain exhibits bright green fluorescence when bound to RNA, but only a weak fluorescent signal when bound to DNA. |
SYTOX dyes are cell-impermeant nucleic acid stains making them useful dead-cell stains. DAPI is a classic nuclear and chromosome counterstain for identifying nuclei and observing chromosome-banding patterns. Upon binding the minor groove of double-stranded DNA, its fluorescence is ~20-fold greater than the unbound state.
Learn more about SYTOX nucleic acid stains
DAPI | SYTOX Blue Nucleic Acid Stain | SYTOX Green Nucleic Acid Stain | SYTOX Orange Nucleic Acid Stain | SYTOX Deep Red Nucleic Acid Stain | TO-PRO | |
---|---|---|---|---|---|---|
Readout | Fluorescent staining of nucleic acids | |||||
Target | Binds double-stranded DNA | |||||
Common filter set | DAPI | Violet/DAPI | FITC | RFP/TRITC | Cy5 | Cy5 |
Labels | DAPI | SYTOX Blue | SYTOX Green | SYTOX Orange | SYTOX Deep Red | TO-PRO-3 |
Ex/Em (nm) | 360/460 | 444/480 | 483/503 | 547/570 | 660/682 | 642/661 |
Signal-to-noise ratio | ||||||
Photostability | ||||||
Multiplexing | Yes | Yes | Yes | Yes | Yes | Yes |
Live cells | Semi-permeant | No | No | No | No | No |
Fixed cells | Yes | Yes | Yes | Yes | Yes | Yes |
Fixable | Yes | Yes | Yes | Yes | Yes | No |
Platform | Imaging | Imaging | Imaging | Imaging | Imaging | Imaging |
Protocol | — | — | Imaging protocol | — | — | Imaging protocol |
Format | 1 mL | 250 μL | 250 µL | 250 µL | 5 x 50 µL | 1 mL |
Cat. No. | 62248 | S11348 | S7020 | S11368 | S11381 | T3605 |
SelectFX Nuclear Labeling Kit includes four spectrally distinct fluorescent dyes for staining nuclei in fixed cells: blue fluorescent DAPI, green-fluorescent SYTOX Green stain, red-fluorescent 7-aminoactinomycin D (7-AAD), and far red-fluorescent TO-PRO-3 dye. |
CellLight fluorescent fusion proteins label histone proteins and can multiplexed with other fluorescent probes in live cells or after fixation.
Learn more about nuclear CellLight reagents
Readout |
Expression of fluorescent fusion protein
| Expression of fluorescent fusion protein | |
Target |
Labels histone proteins within the chromosome
| Localizes fluorescent protein in the nucleus | |
Common filter set | FITC | FITC | TRITC |
Labels | GFP | GFP | RFP |
Ex/Em (nm) | 488/520 | 488/520 | 555/584 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
Yes
|
Fixed cells |
No
|
No
|
No
|
Fixable |
Yes
|
Yes
|
Yes
|
Platform |
Imaging
|
Imaging
|
Imaging
|
Format |
1 mL
|
1 mL
|
1 mL
|
Cat. No. |
HCS NuclearMask stains can be used to measure DNA content and perform cell demarcation on high-content imaging and analysis (HCS) platforms. These stains can be applied to live or fixed cells—able to survive standard formaldehyde-based fixation and detergent-based permeabilized methods. HCS NuclearMask stains are available in a choice of colors for multiplexing flexibility with other functional and structural probes.
Learn more about HCS reagents
Readout |
Stains nucleus for demarcation
| ||
Common filter set | DAPI | Texas Red | Cy5 |
Ex/Em (nm) | 350⁄451 | 622⁄645 | 638⁄686 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
Yes
|
Fixed cells |
Yes
|
Yes
|
No
|
Fixable |
Yes
|
Yes
|
Yes
|
Platform |
HCS
|
HCS
|
HCS
|
Protocol | HCS protocol | HCS protocol | HCS protocol |
Format |
65 µL
|
125 µL
|
400 µL
|
Cat. No. |
ReadyProbes reagents provide the easiest fluorescent staining method for nucleic acids in live or fixed cells. These are ready-to-use solutions in convenient dropper bottles, formulated for room temperature storage.
Learn more about ReadyProbes reagents
Readout |
Fluorescent staining of nucleic acids
| ||
Target |
Membrane-permeable dyes targeting RNA and DNA
|
Membrane impermeable dye targeting RNA and DNA
| |
Common filter set | DAPI | Cy5 | DAPI |
Labels | Hoechst 33342 | NucRed Live | DAPI |
Ex/Em (nm) | 360/460 | 638/686 | 360/460 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
No
|
Fixed cells |
No
|
No
|
Yes
|
Fixable |
Yes
|
No
|
No
|
Platform |
Imaging
|
Imaging
|
Imaging
|
Protocol | Microscopy protocol | Microscopy protocol | Microscopy protocol |
Format |
6 x dropper bottles
|
6 x dropper bottles
|
6 x dropper bottles
|
Cat. No. |
SYTO dyes are live cell, cell-permeant nucleic acid stains that exhibit bright fluorescence upon binding to nucleic acids. Hoechst dyes are blue-fluorescent nucleic acid stains that have multiple applications including distinguishing condensed pycnotic nuclei in apoptotic cells and for cell-cycle studies in combination with BrdU.
Learn more about SYTO nucleic acid stains
Hoechst 33342 | SYTO 9 Green Nucleic Acid Stain | SYTO 82 Orange Nucleic Acid Stain | SYTO 59 Red Nucleic Acid Stain | SYTO Deep Red Nucleic Acid Stain | |
---|---|---|---|---|---|
Readout | Fluorescent staining of nucleic acids | ||||
Target | Preferentially binds AT regions of dsDNA | Cell-permeant dyes targeting DNA and RNA | |||
Common filter set | UV / DAPI | FITC | TRITC | Texas Red | Cy5 |
Labels | Hoechst 33342 | SYTO 9 | SYTO 82 | SYTO 59 | SYTO Deep Red |
Ex/Em (nm) | 350/461 | 483/503 | 541/560 | 622/645 | 652/669 |
Signal-to-noise ratio | |||||
Photostability | |||||
Multiplexing | Yes | Yes | Yes | Yes | Yes |
Live cells | Yes | Yes | Yes | Yes | Yes |
Fixed cells | No | No | No | Yes | No |
Fixable | Yes | Yes | Yes | Yes | Yes |
Platform | Imaging | Imaging | Imaging | Imaging | Imaging |
Protocol | Imaging protocol | — | Imaging protocol | Imaging protocol | — |
Format | 10 mL | 100 μL | 250 µL | 100 µL | 1 vial |
Cat. No. | H3570 | S34854 | S11363 | S11341 | S34900 |
SYTO RNASelect Green Fluorescent Cell Stain is a cell-permeant nucleic acid stain that is selective for RNA. Although virtually nonfluorescent in the absence of nucleic acids, the SYTO RNASelect stain exhibits bright green fluorescence when bound to RNA, but only a weak fluorescent signal when bound to DNA. |
SYTOX dyes are cell-impermeant nucleic acid stains making them useful dead-cell stains. DAPI is a classic nuclear and chromosome counterstain for identifying nuclei and observing chromosome-banding patterns. Upon binding the minor groove of double-stranded DNA, its fluorescence is ~20-fold greater than the unbound state.
Learn more about SYTOX nucleic acid stains
DAPI | SYTOX Blue Nucleic Acid Stain | SYTOX Green Nucleic Acid Stain | SYTOX Orange Nucleic Acid Stain | SYTOX Deep Red Nucleic Acid Stain | TO-PRO | |
---|---|---|---|---|---|---|
Readout | Fluorescent staining of nucleic acids | |||||
Target | Binds double-stranded DNA | |||||
Common filter set | DAPI | Violet/DAPI | FITC | RFP/TRITC | Cy5 | Cy5 |
Labels | DAPI | SYTOX Blue | SYTOX Green | SYTOX Orange | SYTOX Deep Red | TO-PRO-3 |
Ex/Em (nm) | 360/460 | 444/480 | 483/503 | 547/570 | 660/682 | 642/661 |
Signal-to-noise ratio | ||||||
Photostability | ||||||
Multiplexing | Yes | Yes | Yes | Yes | Yes | Yes |
Live cells | Semi-permeant | No | No | No | No | No |
Fixed cells | Yes | Yes | Yes | Yes | Yes | Yes |
Fixable | Yes | Yes | Yes | Yes | Yes | No |
Platform | Imaging | Imaging | Imaging | Imaging | Imaging | Imaging |
Protocol | — | — | Imaging protocol | — | — | Imaging protocol |
Format | 1 mL | 250 μL | 250 µL | 250 µL | 5 x 50 µL | 1 mL |
Cat. No. | 62248 | S11348 | S7020 | S11368 | S11381 | T3605 |
SelectFX Nuclear Labeling Kit includes four spectrally distinct fluorescent dyes for staining nuclei in fixed cells: blue fluorescent DAPI, green-fluorescent SYTOX Green stain, red-fluorescent 7-aminoactinomycin D (7-AAD), and far red-fluorescent TO-PRO-3 dye. |
CellLight fluorescent fusion proteins label histone proteins and can multiplexed with other fluorescent probes in live cells or after fixation.
Learn more about nuclear CellLight reagents
Readout |
Expression of fluorescent fusion protein
| Expression of fluorescent fusion protein | |
Target |
Labels histone proteins within the chromosome
| Localizes fluorescent protein in the nucleus | |
Common filter set | FITC | FITC | TRITC |
Labels | GFP | GFP | RFP |
Ex/Em (nm) | 488/520 | 488/520 | 555/584 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
Yes
|
Fixed cells |
No
|
No
|
No
|
Fixable |
Yes
|
Yes
|
Yes
|
Platform |
Imaging
|
Imaging
|
Imaging
|
Format |
1 mL
|
1 mL
|
1 mL
|
Cat. No. |
HCS NuclearMask stains can be used to measure DNA content and perform cell demarcation on high-content imaging and analysis (HCS) platforms. These stains can be applied to live or fixed cells—able to survive standard formaldehyde-based fixation and detergent-based permeabilized methods. HCS NuclearMask stains are available in a choice of colors for multiplexing flexibility with other functional and structural probes.
Learn more about HCS reagents
Readout |
Stains nucleus for demarcation
| ||
Common filter set | DAPI | Texas Red | Cy5 |
Ex/Em (nm) | 350⁄451 | 622⁄645 | 638⁄686 |
Signal-to-noise ratio | |||
Photostability | |||
Multiplexing |
Yes
|
Yes
|
Yes
|
Live cells |
Yes
|
Yes
|
Yes
|
Fixed cells |
Yes
|
Yes
|
No
|
Fixable |
Yes
|
Yes
|
Yes
|
Platform |
HCS
|
HCS
|
HCS
|
Protocol | HCS protocol | HCS protocol | HCS protocol |
Format |
65 µL
|
125 µL
|
400 µL
|
Cat. No. |
SYTO stains are cell-permeant stains that exhibit bright fluorescence upon binding to nucleic acids in live and dead cells (Figure 1). These dyes can also stain Gram-positive and Gram-negative bacteria. There are a range of wavelengths available, and each stain has a different fluorescence intensity, binding affinity, and nucleic acid selectivity. It is important to note that SYTO dyes do not exclusively stain the nucleus like DNA-selective dyes, such as DAPI or Hoechst 33342. Cells stained with SYTO dyes generally show cytoplasmic or mitochondrial staining.
The nucleolus is a non-membrane-bound structure found within the nucleus and is the site where ribosomal RNA is transcribed. The SYTO RNASelect Green Fluorescent Cell Stain is a cell-permeant nucleic acid stain that is selective for RNA. When bound to RNA, the RNASelect stain will fluoresce bright green but when bound to DNA, it will weakly fluoresce. The SYTO RNASelect stain can be used to detect nucleoli, and cells may be counterstained with a nuclear stain such as DAPI (Figure 2).
SYTO RNASelect Green Fluorescent Cell Stain can be used in live cells or fixed cells. After live cells have been stained, they may be methanol fixed with a minimal loss of staining pattern. Formaldehyde fixation is not recommended since the staining pattern will be altered.
Figure 2. Methanol-fixed MRC-5 cells stained with SYTO RNASelect green-fluorescent cell stain. Methanol-fixed MRC-5 cells stained with SYTO RNASelect green-fluorescent cell stain Nuclei were stained with DAPI; the densely stained areas are nucleoli.
SYTOX nucleic acid stains are cell-impermeant, high-affinity nucleic acid stains that will cross a compromised membrane but will not cross the membrane of live cells. This makes them useful dead-cell stains. Upon binding nucleic acids, there is a >500-fold increase in fluorescence. SYTOX dead cell stains are available in a variety of colors for multiplexing and colocalization studies in fixed cells. They are also incorporated into several assays for apoptosis and cell viability.
SYTOX stains can be used in various applications. For example, SYTOX Blue and SYTOX Green nucleic acid stains have been used for DNA counterstains for chromosome labeling. They can also be used for Gram-negative and Gram-positive bacterial staining. SYTOX Deep Red nucleic acid stain has been used for single-cell analysis (Figure 3), tissue sections, 3D cell models, and bacteria.
CellLight fluorescent fusion proteins are available for nuclear and nuclear protein labeling. CellLight Nucleus (Figure 4) uses a SV40 nuclear localization sequence (1.0 kDa) for identification and demarcation of the nucleus in live-cell imaging experiments. CellLight Histone (Figure 5) uses a histone 2B sequence (13.7 kDa) to follow the dynamics of chromosomal behavior and is particularly useful for real-time imaging of mitotic cell division.
Introducing CellLight fluorescent proteins involves a simple transfection step using the BacMam technology, and they work like cell stains with minimal toxicity or chemical disruption. These nucleus fusion proteins are compatible with other fluorescent probes for multiplex analysis in live cells, or after formaldehyde fixation for colocalization studies.
Learn more about these and other CellLight fluorescent reagents
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