ReadiLink™ iFluor® 488 FISH Fluorescence Imaging Kit

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
International	See distributors
Bulk request	Inquire
Custom size	Inquire
Shipping	Standard overnight for United States, inquire for international

Physical properties

Solvent

Water

Spectral properties

Correction Factor (260 nm)	0.21
Correction Factor (280 nm)	0.11
Extinction coefficient (cm ^-1 M ^-1)	75000¹
Excitation (nm)	491
Emission (nm)	516
Quantum yield	0.9¹

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
Storage	Freeze (< -15 °C); Minimize light exposure

Overview SDS Protocol

Correction Factor (260 nm)

0.21

Correction Factor (280 nm)

0.11

Extinction coefficient (cm ^-1 M ^-1)

75000¹

Excitation (nm)

491

Emission (nm)

516

Quantum yield

0.9¹

Fluorescence in situ hybridization (FISH) technology is an effective tool for detecting specific nucleic acid targets in a biological specimen. Detection of a nucleic acid target in situ is achieved through the hybridization of a fluorescent dye-labeled nucleic acid probe of complementary sequence to the specimen. The Readilink™ iFluor® 488 FISH fluorescence imaging kit is a convenient tool for labeling a target DNA using an iFluor® 488 labeled FISH probe via in situ hybridization. The kit provides Taq DNA polymerase enzyme, which incorporates iFluor® 488-dUTPs in the target DNA through Polymerase Chain Reaction (PCR). Our proprietary iFluor® dyes are brighter and more photostable than traditional fluorescent labels, providing the desired resolution and signal.

Platform

Thermal Cycler

Recommended plate

PCR Microplate

Components

Example protocol

SAMPLE EXPERIMENTAL PROTOCOL

Before using, thaw all components to room temperature and mix thoroughly by vortexing.

Note: The following protocol can be used as a general guideline to standard DNA FISH. Optimization may be necessary for your experimental system.

Prepare the following reaction mixes as indicated in Table 1.

Table 1. Reagents composition per well for each reaction.

Components	Volume (25 µL/reaction)	Final Conc.
FISH Reaction mix (2X)	12.5 µL	1X
Upstream primer, 10 µM	0.25-2.5 µL	0.1-1.0 µM
Downstream primer, 10 µM	0.25-2.5 µL	0.1-1.0 µM
DNA template	1-5 µL	Optimized conc.
iFluor® 488-dUTP	2.5 µL
dNTP mix	1 µL
Water, nuclease-free	25 µL

Carefully mix the reagents by gentle vortexing followed by a brief centrifuge.
Set up the plate in the qPCR instrument and run as indicated in Table 2.

Table 2. Thermal cycling parameters.

Parameter

Polymerase Activation

PCR (30-40 cycles)

Hold

Denature

Anneal

Extend

Temperature

95 °C

95 °C

55-65 °C

68-72 °C

Time (m:ss)

0:20

0:30

1:00

1:00

Spectrum

Open in Advanced Spectrum Viewer

Spectral properties

Correction Factor (260 nm)	0.21
Correction Factor (280 nm)	0.11
Extinction coefficient (cm ^-1 M ^-1)	75000¹
Excitation (nm)	491
Emission (nm)	516
Quantum yield	0.9¹

Product Family

Name	Excitation (nm)	Emission (nm)	Extinction coefficient (cm ^-1 M ^-1)	Quantum yield	Correction Factor (260 nm)	Correction Factor (280 nm)	Correction Factor (656 nm)
ReadiLink™ iFluor® 555 FISH Fluorescence Imaging Kit	557	570	100000¹	0.64¹	0.23	0.14	-
ReadiLink™ iFluor® 647 FISH Fluorescence Imaging Kit	656	670	250000¹	0.25¹	0.03	0.03	0.0793

Images

Figure 1. Telomere quantitative fluorescence in situ hybridization in metaphase HeLa cells using iFluor® 488-dUTP labeled telomere probes. Probes were created using the ReadiLink™ iFluor® 488 FISH Fluorescence Imaging kit.

References

View all 19 references: Citation Explorer

Molecular Characterization of Mesothelioma: Impact of Histologic Type and Site of Origin on Molecular Landscape.
Authors: Dagogo-Jack, Ibiayi and Madison, Russell W and Lennerz, Jochen K and Chen, Kuei-Ting and Hopkins, Julia F and Schrock, Alexa B and Ritterhouse, Lauren L and Lester, Ashley and Wharton, Keith A and Mino-Kenudson, Mari and Danziger, Natalie and Hung, Yin P and Mata, Douglas A and Ross, Jeffrey S
Journal: JCO precision oncology (2022): e2100422

Fluorescence In Situ Hybridization of Small Non-Coding RNAs.
Authors: Vautrot, Valentin and Heckler, Géraud and Aigueperse, Christelle and Behm-Ansmant, Isabelle
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 73-85

Clinicopathologic Features and Response to Therapy of NRG1 Fusion-Driven Lung Cancers: The eNRGy1 Global Multicenter Registry.
Authors: Drilon, Alexander and Duruisseaux, Michael and Han, Ji-Youn and Ito, Masaoki and Falcon, Christina and Yang, Soo-Ryum and Murciano-Goroff, Yonina R and Chen, Haiquan and Okada, Morihito and Molina, Miguel Angel and Wislez, Marie and Brun, Philippe and Dupont, Clarisse and Branden, Eva and Rossi, Giulio and Schrock, Alexa and Ali, Siraj and Gounant, Valérie and Magne, Fanny and Blum, Torsten Gerriet and Schram, Alison M and Monnet, Isabelle and Shih, Jin-Yuan and Sabari, Joshua and Pérol, Maurice and Zhu, Viola W and Nagasaka, Misako and Doebele, Robert and Camidge, D Ross and Arcila, Maria and Ou, Sai-Hong Ignatius and Moro-Sibilot, Denis and Rosell, Rafael and Muscarella, Lucia Anna and Liu, Stephen V and Cadranel, Jacques
Journal: Journal of clinical oncology : official journal of the American Society of Clinical Oncology (2021): 2791-2802

A novel method of amplified fluorescent in situ hybridization for detection of chromosomal microdeletions in B cell lymphoma.
Authors: Mizuno, Yoshimi and Chinen, Yoshiaki and Tsukamoto, Taku and Takimoto-Shimomura, Tomoko and Matsumura-Kimoto, Yayoi and Fujibayashi, Yuto and Kuwahara-Ota, Saeko and Fujino, Takahiro and Nishiyama, Daichi and Shimura, Yuji and Kobayashi, Tsutomu and Horiike, Shigeo and Taniwaki, Masafumi and Kuroda, Junya
Journal: International journal of hematology (2019): 593-602

FISHing Mycobacterium tuberculosis Complex by Use of a rpoB DNA Probe Bait.
Authors: Loukil, Ahmed and Kirtania, Prithwiraj and Bedotto, Marielle and Drancourt, Michel
Journal: Journal of clinical microbiology (2018)

Super-resolution imaging of a 2.5 kb non-repetitive DNA in situ in the nuclear genome using molecular beacon probes.
Authors: Ni, Yanxiang and Cao, Bo and Ma, Tszshan and Niu, Gang and Huo, Yingdong and Huang, Jiandong and Chen, Danni and Liu, Yi and Yu, Bin and Zhang, Michael Q and Niu, Hanben
Journal: eLife (2017)

Detection of Klebsiella. Pneumoniae Infection with an Antisense Oligomer Against its Ribosomal RNA.
Authors: Chen, Ling and Cheng, Dengfeng and Liu, Guozheng and Dou, Shuping and Wang, Yuzhen and Liu, Xinrong and Liu, Yuxia and Rusckowski, Mary
Journal: Molecular imaging and biology (2016): 527-34

Fluorescence in situ hybridization of small non-coding RNAs.
Authors: Vautrot, Valentin and Aigueperse, Christelle and Branlant, Christiane and Behm-Ansmant, Isabelle
Journal: Methods in molecular biology (Clifton, N.J.) (2015): 73-83

[Application of flow cytometry-fluorescence in situ hybridization in the measurement of relative telomere length of bone marrow CD34(+) cells in patients with myelodysplastic syndrome].
Authors: Guo, Tian-Jiao and Sun, Wan-Ling and Zhang, Wei and Ma, Xiao-Cai and Liu, Cong-Yan and He, Jing-Juan and Xu, Juan
Journal: Zhongguo shi yan xue ye xue za zhi (2013): 1195-9

(99m)Tc-MORF oligomers specific for bacterial ribosomal RNA as potential specific infection imaging agents.
Authors: Chen, Ling and Wang, Yi and Cheng, Dengfeng and Liu, Xinrong and Dou, Shuping and Liu, Guozheng and Hnatowich, Donald J and Rusckowski, Mary
Journal: Bioorganic & medicinal chemistry (2013): 6523-30

Documents

Safety data sheet
Product protocol
Certificate of analysis

Application notes

A Novel Fluorescent Probe for Imaging and Detecting Hydroxyl Radical in Living Cells
Abbreviation of Common Chemical Compounds Related to Peptides
Annexin V
Bright Tide Fluor™-Based Fluorescent Peptides and Their Applications In Drug Discovery and Disease Diagnosis
Calcein

FAQ

Are Cell Navigator® Cell Plasma Membrane Staining Kits suitable for cell culture medium samples?
Are mRNAs found in prokaryotes differ from those of eukaryotes?
Are spliceosomes associated with any diseases?
Are there any alternatives for ethidium bromide in agarose gels?
Are there any alternatives to Cy5?

AssayWise

Glycerol 3-Phosphate Assays
Nucleic Acid Quantification
Hydroxyl Radical Detection
Thiol Detection
Nucleic Acid Detection, Quantification and Imaging

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Parameter	Polymerase Activation	PCR (30-40 cycles)
	Hold	Denature	Anneal	Extend
Temperature	95 °C	95 °C	55-65 °C	68-72 °C
Time (m:ss)	0:20	0:30	1:00	1:00

ReadiLink™ iFluor® 488 FISH Fluorescence Imaging Kit

OverviewSDSProtocol

Platform

Thermal Cycler

Components

Example protocol

SAMPLE EXPERIMENTAL PROTOCOL

Spectrum

Spectral properties

Product Family

Images

References

Documents

Application notes

FAQ

AssayWise

Overview SDS Protocol