AAT Bioquest

Amine Reactive Dyes and Probes for Conjugation

Amine-reactive probes, composed of an amine-reactive moiety and a chemical moiety or protein, are widely used in bioconjugation for their convenience and performance. The strength and stability of the chemical bond between tag and biomolecule produces robust bioconjugates resistant to the harsh conditions of post-processing steps synonymous with applications such as immunohistochemistry (IHC), immunocytochemistry (ICC), flow cytometry, enzyme-linked immunosorbent assays (ELISA), fluorescence in situ hybridization (FISH), cell tracing, and receptor labeling. AAT Bioquest offers a wide variety of amine-reactive probes for labeling proteins, peptides, amine-modified oligonucleotides, and other biomolecules with fluorophores, enzymes, or hapten tags. Featured in this portfolio are our bright and photostable iFluor™ dyes that span the entire UV, visible, and IR spectrum, providing researchers with more possibilities for multiplex detection.

Too busy to perform your conjugation or having difficulty finding the right amine-reactive probe. Not to worry, AAT Bioquest's bioconjugation and custom oligonucleotide labeling services can prepare the bioconjugate you need.




Common Amine-Reactive Moieties and Their Chemical Reactivity

Primary amines (-NH2) are mainly found at the N-terminus of polypeptide chains (i.e., α-amine) and on the side chains of lysine residues (i.e., the ε-amine). Compared to other conjugation targets, such as sulfhydryls (-SH), primary amines have an outward-facing orientation at physiological conditions making them readily accessible for conjugation. When conjugating with amine-reactive probes, it is important to avoid using buffers containing free amines, such as Tris and glycine, as they will compete with the labeling reaction. In addition, any inorganic salts, such as ammonium sulfate, and stabilizing reagents, such as bovine serum albumin (BSA), must be removed via dialysis or some other purification method before labeling. Failure to do so can significantly impact conjugation yield.

While several types of amine-reactive moieties exist, the most commonly used include succinimidyl esters, imidioesters, sulfonyl chlorides, and isothiocyanates. These acylating reagents react with nucleophiles to form acylated products, and as a result, the reactive moiety is displaced. Two key factors to consider when selecting an amine-reactive probe are the reactivity class of the amine group - aliphatic or aromatic - and its basicity. Aliphatic amines, which are located on the ε-amino group of lysine residues, exist on most proteins. They are moderately basic and will react with most acylating reagents optimally at a pH of 8.0 to 9.5. Aromatic amines, however, are not so common. These weak bases are unprotonated at a neutral pH and typically require a highly reactive moiety, such as an isothiocyanate or sulfonly chloride.

Succinimidyl Esters

Succinimidyl ester labeling mechanism
Figure 1. The reaction of a succinimidyl ester compound with a primary amine.
Succinimidyl ester (SE or NHS esters) derivatives are among the most popular conjugation reagents for amine modification. They are relatively easy to prepare, stable for storage, functional in an aqueous environment, and highly reactive for aliphatic amines versus aromatic amines, alcohols, phenols, and histidines. In SE conjugation, the carbonyl carbon of the SE moiety is attacked by a primary aliphatic amine (i.e., nucleophilic acyl substitution), resulting in a tetrahedral intermediate and the displacement of the SE group (i.e., leaving group). The nucleophile takes the place of the leaving group, and a stable amide bond is formed.

SE conjugation is best performed under physiological to slightly alkaline conditions (pH 7-9) with phosphate, carbonate-bicarbonate, HEPES, or borate buffers. Avoid buffers containing primary amines, such as Tris or glycine buffers. These buffers are incompatible with SE reagents and will compete for reaction decreasing conjugation yield. At pH levels closer to 9, the reaction efficiency can improve due to a higher degree of amine deprotonation; however, the rate of SE hydrolysis will also increase. For less concentrated protein solutions, this can result in less-efficient crosslinking.

AAT Bioquest offers a wide selection of SE reagents, including fluorescent and non-fluorescent dye derivatives. These include iFluor® dyes, Alexa Fluor® dye equivalents, conventional fluorophores, non-fluorescent quenchers, pH-sensitive dyes, calcium indicator Cal-520®, biotin, digoxigenin (DIG), DNP, horseradish peroxidase (HRP), estradiol, fentanyl, and more (see table 1 below).

Table 1. Fluorophore, hapten, enzyme and other succinimidyl ester derivatives for labeling proteins, antibodies and peptides.

Ex (nm)
Em (nm)
CF at 280³
Unit Size
Cat No.
5(6)-FAM, SE [5-(and-6)-Carboxyfluorescein, succinimidyl ester] *CAS 117548-22-8*493517830000.17825 mg110
5(6)-FAM, SE [5-(and-6)-Carboxyfluorescein, succinimidyl ester] *CAS 117548-22-8*493517830000.178100 mg111
5(6)-FAM, SE [5-(and-6)-Carboxyfluorescein, succinimidyl ester] *CAS 117548-22-8*493517830000.1781 g112
5-FAM, SE [5-Carboxyfluorescein, succinimidyl ester] *CAS 92557-80-7*493517830000.17810 mg113
5-FAM, SE [5-Carboxyfluorescein, succinimidyl ester] *CAS 92557-80-7*493517830000.178100 mg114
5-FAM, SE [5-Carboxyfluorescein, succinimidyl ester] *CAS 92557-80-7*493517830000.1781 g115
6-FAM, SE [6-Carboxyfluorescein, succinimidyl ester] *CAS 92557-81-8*493517830000.17810 mg116
6-FAM, SE [6-Carboxyfluorescein, succinimidyl ester] *CAS 92557-81-8*493517830000.178100 mg117
6-FAM, SE [6-Carboxyfluorescein, succinimidyl ester] *CAS 92557-81-8*493517830000.1781 g118
Cyanine 3 bissuccinimidyl ester [equivalent to Cy3® bisNHS ester]5555691500000.150.0731 mg138
Cyanine 3 monosuccinimidyl ester [equivalent to Cy3® NHS ester]5555691500000.150.0731 mg141
Cyanine 3.5 monosuccinimidyl ester [equivalent to Cy3.5® NHS ester]5795911500000.150.1781 mg148
Cyanine 5 monosuccinimidyl ester [equivalent to Cy5® NHS ester]6516702500000.270.031 mg151
Cyanine 5 bissuccinimidyl ester [equivalent to Cy5® bisNHS ester]6516702500000.270.031 mg157
Cyanine 5.5 bissuccinimidyl ester [equivalent to Cy5.5® bisNHS ester]6837032500000.270.1011 mg158
Cyanine 7 monosuccinimidyl ester [equivalent to Cy7® NHS ester]7567792500000.30.0361 mg161
Cyanine 7 bissuccinimidyl ester [equivalent to Cy7® bisNHS ester]7567792500000.30.0361 mg170
Cyanine 5.5 monosuccinimidyl ester [equivalent to Cy5.5® NHS ester]6837032500000.270.1011 mg174
Cy3NS succinimidyl ester5555691500000.150.07325 mg191
Cy5NS succinimidyl ester6516702500000.270.0325 mg195
Cy7NS succinimidyl ester7567792500000.30.03625 mg198
Cy5.5NS succinimidyl ester6837032500000.270.10110 mg199
6-HEX, SE [6-Carboxy-2',4,4',5',7,7'-hexachlorofluorescein, succinimidyl ester] *Single Isomer*533559730000.155 mg202
6-JOE, SE [6-Carboxy-4',5'-dichloro-2',7'-dimethoxyfluorescein, succinimidyl ester] *CAS#: 113394-23-3*520545750005 mg203
6-TET, SE [6-Carboxy-2',4,7',7-tetrachlorofluorescein, succinimidyl ester] 521542730000.135 mg211
Helix Fluor™ 545, succinimidyl ester5265431 mg250
Helix Fluor™ 575, succinimidyl ester5535701 mg251
Cyanine 3 bissuccinimidyl ester, potassium salt [same as GE Cy3® bisNHS ester]5555691500000.150.0731 mg270
Cyanine 3 monosuccinimidyl ester, potassium salt [same as GE Cy3® NHS ester]5555691500000.150.0731 mg271
Cyanine 3.5 monosuccinimidyl ester, potassium salt [same as GE Cy3.5® NHS ester]5795911500000.150.1781 mg275
Cyanine 5 monosuccinimidyl ester, potassium salt [same as GE Cy5® NHS ester]6516702500000.270.031 mg280
Cyanine 5 bissuccinimidyl ester, potassium salt [same as GE Cy5® bisNHS ester]6516702500000.270.031 mg282
Cyanine 5.5 monosuccinimidyl ester, potassium salt [same as Cy5.5® NHS ester]6837032500000.270.1011 mg283
Cyanine 5.5 bissuccinimidyl ester, potassium salt [same as GE Cy5.5® bisNHS ester]6837032500000.270.1011 mg285
Cyanine 7 monosuccinimidyl ester, potassium salt [same as GE Cy7® NHS ester]7567792500000.30.0361 mg290
Cyanine 7 bissuccinimidyl ester, potassium salt [same as GE Cy7® bisNHS ester]7567792500000.30.0361 mg295
5(6)-CR6G, SE [5-(and 6)-Carboxyrhodamine 6G, succinimidyl ester] *Mixed isomers*522546940000.21410 mg340
5-CR6G, SE [5-Carboxyrhodamine 6G, succinimidyl ester] *CAS 209112-21-0*522546940000.2141 mg341
6-CR6G, SE [6-Carboxyrhodamine 6G, succinimidyl ester] *Single isomer*522546940000.2141 mg342
6-CR6G, SE [6-Carboxyrhodamine 6G, succinimidyl ester] *Single isomer*522546940000.2145 mg346
5-CR6G, SE [5-Carboxyrhodamine 6G, succinimidyl ester] *Single isomer*522546940000.2145 mg345
5(6)-TAMRA, SE [5-(and-6)-Carboxytetramethylrhodamine, succinimidyl ester] *CAS 246256-50-8*552578900000.17825 mg370
5(6)-TAMRA, SE [5-(and-6)-Carboxytetramethylrhodamine, succinimidyl ester] *CAS 246256-50-8*552578900000.178100 mg371
5(6)-TAMRA, SE [5-(and-6)-Carboxytetramethylrhodamine, succinimidyl ester] *CAS 246256-50-8*552578900000.1781 g372
5-TAMRA, SE [5-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-68-7*552578900000.1785 mg373
5-TAMRA, SE [5-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-68-7*552578900000.178100 mg374
5-TAMRA, SE [5-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-68-7*552578900000.1781 g375
6-TAMRA, SE [6-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-69-8*552578900000.1785 mg376
6-TAMRA, SE [6-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-69-8*552578900000.178100 mg377
6-TAMRA, SE [6-Carboxytetramethylrhodamine, succinimidyl ester] *CAS#: 150810-69-8*552578900000.1781 g378
5(6)-ROX, SE [5-(and-6)-Carboxy-X-rhodamine, succinimidyl ester] *Mixed isomers*578604820000.16825 mg390
5-ROX, SE [5-Carboxy-X-rhodamine, succinimidyl ester] *CAS 209734-74-7*578604820000.1685 mg389
5-ROX, SE [5-Carboxy-X-rhodamine, succinimidyl ester] *CAS 209734-74-7*578604820000.168100 mg391
6-ROX, SE [6-Carboxy-X-rhodamine, succinimidyl ester] *CAS#: 216699-36-4*578604820000.1685 mg392
6-ROX, SE [6-Carboxy-X-rhodamine, succinimidyl ester] *CAS#: 216699-36-4*578604820000.168100 mg399
6-ROX-AHA, SE578604820000.1685 mg393
LRB Red™ SE5585750.17110 mg471
SunRed™ SE592609-0.3665 mg472
SunRed™ SE5926090.3661 mg478
California Red™ SE5926091000000.1945 mg473
California Red™ SE5926091000000.1941 mg479
Texas Red-X, succinimidyl ester *Single isomer* *178623-11-5*5866031160000.930.365 mg474
AMCA, succinimidyl ester346434190000.15310 mg502
DEAC,SE [7-Diethylaminocoumarin-3-carboxylic acid, succinimidyl ester] *CAS 139346-57-9*4114720.1225 mg506
7-Hydroxycoumarin-3-carboxylic acid, succinimidyl ester *CAS 134471-24-2*3524070.081550 mg551
7-Hydroxycoumarin-4-acetic acid, succinimidyl ester25 mg553
7-Hydroxy-4-methylcoumarin-3-acetic acid, succinimidyl ester25 mg556
MCA succinimidyl ester [7-Methoxycoumarin-4-acetic acid, succinimidyl ester]3223810.325 mg558
7-Methoxycoumarin-3-carboxylic acid, succinimidyl ester *CAS 150321-92-9*3554050.171100 mg563
Bodi Fluor™ R6G NHS Ester [equivalent to Bodipy® R6G, NHS Ester]5285471160005 mg703
Dansyl-X, SE33551833000.3871 g813
NBD-X, succinimidyl ester *CAS 145195-58-0*46753922000100 mg829
iFluor® A7 SE7627822750000.10.031 mg1039
iFluor® A7 SE7627822750000.10.03100 ug71039
iFluor® A7 SE7627822750000.10.035 mg71517
iFluor® A7 SE7627822750000.10.0310 mg71567
mFluor™ Violet 450 SE406445350000.810.0781 mg1150
mFluor™ Violet 510 SE412505250000.860.3661 mg1151
mFluor™ Violet 540 SE402535180000.210.5431 mg1152
mFluor™ Blue 570 SE5535651200000.1791 mg1160
mFluor™ Green 620 SE525623500000.5691 mg1165
mFluor™ Yellow 630 SE5706321100001 mg1170
mFluor™ Red 700 SE6806952950000.1271 mg1190
mFluor™ Red 780 SE629767900000.1161 mg1191
trFluor™ Eu succinimidyl ester3466170.7771 mg1433
trFluor™ Tb succinimidyl ester3335440.7971 mg1443
DABCYL succinimidyl ester [4-((4-(Dimethylamino)phenyl)azo)benzoic acid, succinimidyl ester] *CAS 146998-31-4*0.5161 g2004
DABCYL succinimidyl ester [4-((4-(Dimethylamino)phenyl)azo)benzoic acid, succinimidyl ester] *CAS 146998-31-4*0.5165 g2005
DNP-X acid, SE [6-(2,4-Dinitrophenyl)aminohexanoic acid, succinimidyl ester] *CAS 82321-04-8*25 mg2021
Tide Quencher™ 2WS succinimidyl ester [TQ2WS, SE]480000.1205 mg2058
Tide Quencher™ 4WS succinimidyl ester [TQ4WS SE]900000.1361 mg2067
Tide Quencher™ 5WS succinimidyl ester [TQ5WS SE]1300000.0821 mg2081
Tide Quencher™ 6WS succinimidyl ester [TQ6WS SE]1300000.1021 mg2096
Tide Quencher™ 7WS succinimidyl ester [TQ7WS SE]1400000.0911 mg2111
Tide Quencher™ 1 succinimidyl ester [TQ1 SE]200000.19425 mg2199
Tide Quencher™ 2 succinimidyl ester [TQ2 SE]210000.1225 mg2210
Tide Quencher™ 3WS succinimidyl ester [TQ3WS SE]900000.2051 mg2229
Tide Quencher™ 3 succinimidyl ester [TQ3 SE]220000.09125 mg2230
Tide Fluor™ 1 succinimidyl ester [TF1 SE]*Superior replacement for EDANS*341448200000.1875 mg2244
Tide Fluor™ 2, succinimidyl ester [TF2 SE]*Superior replacement for fluorescein*503525750000.90.095 mg2248
Tide Fluor™ 3 succinimidyl ester [TF3 SE]*Superior replacement for Cy3*554578750000.1795 mg2271
Tide Fluor™ 5WS succinimidyl ester [TF5WS SE]*Superior replacement for Cy5*6496642500000.0275 mg2281
Tide Fluor™ 4, succinimidyl ester [TF4 SE]*Superior replacement for ROX and Texas Red*578602900000.4365 mg2289
Tide Fluor™ 6WS succinimidyl ester [TF6WS SE]*Superior replacement for Cy5.5*6827012200000.1011 mg2294
Tide Fluor™ 7WS, succinimidyl ester [TF7WS SE]*Superior replacement for Cy7*7567802750000.0491 mg2333
Tide Fluor™ 8WS, succinimidyl ester [TF8WS SE]*Near Infrared Emission*7858012500000.1091 mg2338
Tide Fluor™ 3WS succinimidyl ester [TF3WS SE] *Superior replacement for Cy3*5515631500000.0795 mg2346
Tide Fluor™ 2WS succinimidyl ester [TF2WS SE] *Superior replacement for FITC*491516750000.90.115 mg2349
Biotin, succinimidyl ester *CAS 35013-72-0*100 mg3002
Biotin-X, succinimidyl ester *CAS 72040-63-2*25 mg3010
Biotin PEG2 succinimidyl ester25 mg3016
Biotin PEG4 succinimidyl ester25 mg3022
ReadiView™ biotin succinimidyl ester5 mg3059
ReadiUse™ Preactivated HRP NHS ester300 ug11025
Cal-520® NHS Ester4935150.75100 ug20609
RatioWorks™ BCFL, SE5045271 mg21191
Protonex™ Red 600, SE5765971 mg21208
Protonex™ Green 500, SE44550340001 mg21216
CFSE [5-(and 6)-Carboxyfluorescein diacetate, succinimidyl ester] *CAS 150347-59-4*49851725 mg22022
CDCFDA, SE [5-(and-6)-Carboxy-2',7'-dichlorofluorescein diacetate, succinimidyl ester] *Mixed isomers*50552625 mg22026
ReadiUse™ CFSE [5-(and 6)-Carboxyfluorescein diacetate, succinimidyl ester] *CAS 150347-59-4*4985175x500 ug22028
5-OG488 succinimidyl ester [equivalent to Oregon Green® 488 carboxylic acid, succinimidyl ester, 5-isomer ]498526760000.125 mg710
6-OG488 succinimidyl ester [equivalent to Oregon Green® 488 carboxylic acid, succinimidyl ester, 6-isomer ]498526760000.125 mg711
Texas Red-X, succinimidyl ester *Mixed isomers* *CAS 216972-99-5*5866031160000.930.365 mg475
Digoxigenin NHS ester1 mg3500
Digoxigenin NHS ester5 mg3501
PacBlue succinimidyl ester [equivalent to Pacific Blue succinimidyl ester]404455460000.780.25 mg570
zFluor™ 647 succinimidyl ester6486681 mg1500
zFluor™ 633 succinimidyl ester6236391 mg1510
XFD488 NHS Ester *Same Structure to Alexa Fluor™ 488 NHS Ester*499520730000.921 mg1812
XFD488 NHS Ester *Same Structure to Alexa Fluor™ 488 NHS Ester*499520730000.925 mg71902
XFD350 NHS Ester *Same Structure to Alexa Fluor™ 350 NHS Ester*343441190001 mg1800
XFD350 NHS Ester *Same Structure to Alexa Fluor™ 350 NHS Ester*343441190005 mg71900
XFD532 NHS Ester *Same Structure to Alexa Fluor™ 532 NHS Ester*534553810000.611 mg1819
XFD532 NHS Ester *Same Structure to Alexa Fluor™ 532 NHS Ester*534553810000.615 mg71904
XFD594 NHS Ester *Same Structure to Alexa Fluor™ 594 NHS Ester*590618920000.661 mg1830
XFD594 NHS Ester *Same Structure to Alexa Fluor™ 594 NHS Ester*590618920000.665 mg71908
Thioflavin T, succinimidyl ester3494545 mg23065
mFluor™ Violet 500 SE410501250000.810.3651 mg1149
Sulfo-Cyanine 3 NHS ester5555691500000.150.07310 mg263
Sulfo-Cyanine 3 NHS ester5555691500000.150.07350 mg264
Sulfo-Cyanine 5 NHS ester6516702500000.270.0310 mg268
Sulfo-Cyanine 5 NHS ester6516702500000.270.0350 mg269
RuFluor™ succinimidyl ester1 mg1520
trFluor™ Eu-Cryptate succinimidyl ester30561728000100 ug1430
trFluor™ Eu-Cryptate succinimidyl ester305617280001 mg1431
6-VIC, SE [6-VIC NHS ester]5265431 mg212
6-VIC, SE [6-VIC NHS ester]5265435 mg213
Lumiwox™ acridinium NHS ester1 mg26000
mFluor™ UV375 SE351387300000.940.1381 mg1135
XFD488 NHS Ester-UltraPure Grade *XFD488 Same Structure to Alexa Fluor™ 488*499520730000.921 mg1810
XFD488 NHS Ester-UltraPure Grade *XFD488 Same Structure to Alexa Fluor™ 488*499520730000.925 mg71901
QXY21 NHS ester  [equivalent to QSY-21 NHS ester]890000.3210 mg2033
XFD555 NHS Ester *Same Structure to Alexa Fluor™ 555 NHS Ester*5535681550000.11 mg1821
XFD555 NHS Ester *Same Structure to Alexa Fluor™ 555 NHS Ester*5535681550000.15 mg71906
XFD647 NHS Ester *Same Structure to Alexa Fluor™ 647 NHS Ester*6506712700000.331 mg1833
XFD647 NHS Ester *Same Structure to Alexa Fluor™ 647 NHS Ester*6506712700000.335 mg71909
XFD680 NHS Ester *Same Structure to Alexa Fluor™ 680 NHS Ester*6817041320000.361 mg1835
XFD680 NHS Ester *Same Structure to Alexa Fluor™ 680 NHS Ester*6817041320000.365 mg71910
XFD700 NHS Ester *Same Structure to Alexa Fluor™ 700 NHS Ester*6967192050000.251 mg1836
XFD700 NHS Ester *Same Structure to Alexa Fluor™ 700 NHS Ester*6967192050000.255 mg71911
XFD750 NHS Ester *Same Structure to Alexa Fluor™ 750 NHS Ester*7527762900000.121 mg1839
XFD750 NHS Ester *Same Structure to Alexa Fluor™ 750 NHS Ester*7527762900000.125 mg71912
Buccutite™ MTA, NHS ester2 umoles5355
N-Me-DMAE, NHS ester5 mg26005
N-Sp-DMAE, NHS ester5 mg26010
Acridinium C2, NHS Ester5 mg26015
Cy5DIGE NHS ester6516702500000.270.03100 nmoles25306
Cy2DIGE NHS ester4925081500000.120.15100 nmoles25300
Cy3DIGE NHS ester5555691500000.150.073100 nmoles25303
DBCO NHS Ester10 mg4529
DBCO-PEG4-NHS Ester1 mg4530
mFluor™ UV460 SE358456150000.860.1341 mg1136
cAMP-NHS ester1 mg17102
Alkyne Modifier I, NHS ester [PEG1 alkyne NHS ester]25 mg4531
Alkyne Modifier II, NHS ester [PEG4 alkyne NHS ester]25 mg4533
XFD546 NHS Ester *Same Structure to Alexa Fluor™ 546 NHS Ester*5615721120000.791 mg1820
XFD546 NHS Ester *Same Structure to Alexa Fluor™ 546 NHS Ester*5615721120000.795 mg71905
XFD568 NHS Ester *Same Structure to Alexa Fluor™ 568 NHS Ester*579603880000.691 mg1823
XFD568 NHS Ester *Same Structure to Alexa Fluor™ 568 NHS Ester*579603880000.695 mg71907
RatioWorks™ PH165, NHS ester4636411 mg21214
AzoDye-1 Succinimidyl Ester5 mg2402
AzoDye-2 Succinimidyl Ester5 mg2422
5-dR6G, succinimidyl ester5 mg302
5-dTMR, succinimidyl ester5 mg307
5-dROX, succinimidyl ester5 mg312
5-dR110, succinimidyl ester5 mg317
Estradiol succinimidyl ester1 mg50545
Fentanyl succinimidyl ester1 mg50560
CDPI3 Succinimidyl Ester [Minor Groove Binder SE]1 mg6907
mFluor™ Blue 630 SE470634490000.2751 mg1164
mFluor™ Blue 660 SE481663260000.321 mg1180
Bodi Fluor™ 576/589 NHS Ester (equivalent to BODIPY™ 576/589 NHS Ester)5735925 mg661
mFluor™ Violet 610 SE421613350000.30.661 mg1156
BG Succinimidyl Ester1 mg12564
BC Succinimidyl Ester1 mg12565
mFluor™ Green 630 SE537657510001 mg1168
mFluor™ Blue 580 SE485580400000.2471 mg1178
6-NED, SE [6-NED NHS ester]545567740001 mg214
6-NED, SE [6-NED NHS ester]545567740005 mg215
mFluor™ Blue 590 SE569589810000.150.4061 mg1161
mFluor™ Blue 620 SE589616980000.180.8491 mg1163
DNP-PEG4 NHS ester5 mg2023
zFluor™ 635 succinimidyl ester667692110,0000.220.091 mg1501
NTA succinimidyl ester5 mg12608
mFluor™ Violet 550 SE527550900000.310.3061 mg1153
mFluor™ Violet 505 SE393504400000.450.4031 mg1154
mFluor™ Violet 590 SE564591900000.220.3291 mg1155
mFluor™ Violet 545 SE393543200000.150.4961 mg1157
mFluor™ UV420 SE353421750001 mg1641
mFluor™ UV455 SE357461200000.420.4061 mg1642
mFluor™ UV520 SE503524800000.5181 mg1643
mFluor™ UV540 SE542560900000.350.4631 mg1645
mFluor™ UV610 SE590609900000.250.9041 mg1649
XFD514 NHS Ester *Same Structure to Alexa Fluor™ 514 NHS Ester*518543800001 mg1818
XFD514 NHS Ester *Same Structure to Alexa Fluor™ 514 NHS Ester*518543800005 mg71903
QXY7 acid  [equivalent to QSY-7 NHS ester]9000025 mg2037
QXY7 NHS ester  [equivalent to QSY-7 NHS ester]9000010 mg2038
IDA succinimidyl ester5 mg12632
RatioWorks™ PDMPO, SE3335311 mg21210
Buccutite™ FOL, NHS ester2 umoles5350
Cy3B NHS ester5605711200000.580.0691 mg941
Cy3B NHS ester5605711200000.580.0695 mg949
iFluor® 350 succinimidyl ester345450200000.950.231 mg1020
iFluor® 350 succinimidyl ester345450200000.950.23100 ug71020
iFluor® 350 succinimidyl ester345450200000.950.235 mg71500
iFluor® 350 succinimidyl ester345450200000.950.2310 mg71550
iFluor® 405 succinimidyl ester403427370000.910.771 mg1021
iFluor® 405 succinimidyl ester403427370000.910.77100 ug71021
iFluor® 405 succinimidyl ester403427370000.910.775 mg71501
iFluor® 405 succinimidyl ester403427370000.910.7710 mg71551
iFluor® 488 succinimidyl ester491516750000.90.111 mg1023
iFluor® 488 succinimidyl ester491516750000.90.11100 ug71023
iFluor® 488 succinimidyl ester491516750000.90.115 mg71503
iFluor® 488 succinimidyl ester491516750000.90.1110 mg71553
iFluor® 514 succinimidyl ester511527750000.830.1161 mg1024
iFluor® 514 succinimidyl ester511527750000.830.116100 ug71024
iFluor® 514 succinimidyl ester511527750000.830.1165 mg71504
iFluor® 514 succinimidyl ester511527750000.830.11610 mg71554
iFluor® 532 succinimidyl ester537560900000.680.161 mg1025
iFluor® 532 succinimidyl ester537560900000.680.16100 ug71025
iFluor® 532 succinimidyl ester537560900000.680.165 mg71505
iFluor® 532 succinimidyl ester537560900000.680.1610 mg71555
iFluor® 555 succinimidyl ester5575701000000.640.141 mg1028
iFluor® 555 succinimidyl ester5575701000000.640.14100 ug71028
iFluor® 555 succinimidyl ester5575701000000.640.145 mg71507
iFluor® 555 succinimidyl ester5575701000000.640.1410 mg71557
iFluor® 594 succinimidyl ester5886041800000.530.041 mg1029
iFluor® 594 succinimidyl ester5886041800000.530.04100 ug71029
iFluor® 594 succinimidyl ester5886041800000.530.045 mg71508
iFluor® 594 succinimidyl ester5886041800000.530.0410 mg71558
iFluor® 633 succinimidyl ester6406542500000.290.0441 mg1030
iFluor® 633 succinimidyl ester6406542500000.290.044100 ug71030
iFluor® 633 succinimidyl ester6406542500000.290.0445 mg71509
iFluor® 633 succinimidyl ester6406542500000.290.04410 mg71559
iFluor® 647 succinimidyl ester6566702500000.250.031 mg1031
iFluor® 647 succinimidyl ester6566702500000.250.03100 ug71031
iFluor® 647 succinimidyl ester6566702500000.250.035 mg71510
iFluor® 647 succinimidyl ester6566702500000.250.0310 mg71560
iFluor® 660 succinimidyl ester6636782500000.260.081 mg1032
iFluor® 660 succinimidyl ester6636782500000.260.08100 ug71032
iFluor® 660 succinimidyl ester6636782500000.260.085 mg71511
iFluor® 660 succinimidyl ester6636782500000.260.0810 mg71561
iFluor® 680 succinimidyl ester6847012200000.230.0941 mg1035
iFluor® 680 succinimidyl ester6847012200000.230.094100 ug71035
iFluor® 680 succinimidyl ester6847012200000.230.0945 mg71513
iFluor® 680 succinimidyl ester6847012200000.230.09410 mg71563
iFluor® 700 succinimidyl ester6907132200000.230.041 mg1036
iFluor® 700 succinimidyl ester6907132200000.230.04100 ug71036
iFluor® 700 succinimidyl ester6907132200000.230.045 mg71514
iFluor® 700 succinimidyl ester6907132200000.230.0410 mg71564
iFluor® 750 succinimidyl ester7577792750000.120.0391 mg1037
iFluor® 750 succinimidyl ester7577792750000.120.039100 ug71037
iFluor® 750 succinimidyl ester7577792750000.120.0395 mg71515
iFluor® 750 succinimidyl ester7577792750000.120.03910 mg71565
iFluor® 610 succinimidyl ester6106281100000.850.491 mg1038
iFluor® 610 succinimidyl ester6106281100000.850.49100 ug71038
iFluor® 610 succinimidyl ester6106281100000.850.495 mg71516
iFluor® 610 succinimidyl ester6106281100000.850.4910 mg71566
iFluor® 710 succinimidyl ester7177391900000.60.071 mg1045
iFluor® 710 succinimidyl ester7177391900000.60.07100 ug71045
iFluor® 710 succinimidyl ester7177391900000.60.075 mg71520
iFluor® 710 succinimidyl ester7177391900000.60.0710 mg71570
iFluor® 790 succinimidyl ester7878122500000.130.091 mg1368
iFluor® 790 succinimidyl ester7878122500000.130.09100 ug71368
iFluor® 790 succinimidyl ester7878122500000.130.095 mg71524
iFluor® 790 succinimidyl ester7878122500000.130.0910 mg71574
iFluor® 800 succinimidyl ester8018202500000.110.081 mg1379
iFluor® 800 succinimidyl ester8018202500000.110.08100 ug71379
iFluor® 800 succinimidyl ester8018202500000.110.085 mg71525
iFluor® 800 succinimidyl ester8018202500000.110.0810 mg71575
iFluor® 810 succinimidyl ester8118222500000.050.151 mg1389
iFluor® 810 succinimidyl ester8118222500000.050.15100 ug71389
iFluor® 810 succinimidyl ester8118222500000.050.155 mg71526
iFluor® 810 succinimidyl ester8118222500000.050.1510 mg71576
iFluor® 820 succinimidyl ester822850250000 0.161 mg1399
iFluor® 820 succinimidyl ester822850250000 0.16100 ug71399
iFluor® 820 succinimidyl ester822850250000 0.165 mg71527
iFluor® 820 succinimidyl ester822850250000 0.1610 mg71577
iFluor® 860 succinimidyl ester853878250000 0.141 mg1409
iFluor® 860 succinimidyl ester853878250000 0.14100 ug71409
iFluor® 860 succinimidyl ester853878250000 0.145 mg71529
iFluor® 860 succinimidyl ester853878250000 0.1410 mg71579
iFluor® 546 succinimidyl ester5415571000000.670.151 mg1048
iFluor® 546 succinimidyl ester5415571000000.670.15100 ug71048
iFluor® 546 succinimidyl ester5415571000000.670.155 mg71521
iFluor® 546 succinimidyl ester5415571000000.670.1510 mg71571
iFluor® 568 succinimidyl ester5685871000000.570.151 mg1049
iFluor® 568 succinimidyl ester5685871000000.570.15100 ug71049
iFluor® 568 succinimidyl ester5685871000000.570.155 mg71522
iFluor® 568 succinimidyl ester5685871000000.570.1510 mg71572
iFluor® 430 succinimidyl ester433498400000.780.31 mg1052
iFluor® 430 succinimidyl ester433498400000.780.3100 ug71052
iFluor® 430 succinimidyl ester433498400000.780.35 mg71523
iFluor® 430 succinimidyl ester433498400000.780.310 mg71573
iFluor® 450 succinimidyl ester451502400000.820.271 mg1026
iFluor® 450 succinimidyl ester451502400000.820.27100 ug71026
iFluor® 450 succinimidyl ester451502400000.820.275 mg71506
iFluor® 450 succinimidyl ester451502400000.820.2710 mg71556
iFluor® 840 succinimidyl ester8368792000000.091 mg1403
iFluor® 840 succinimidyl ester8368792000000.09100 ug71403
iFluor® 840 succinimidyl ester8368792000000.095 mg71528
iFluor® 840 succinimidyl ester8368792000000.0910 mg71578
iFluor® 560 succinimidyl ester5605711200000.570.0691 mg1040
iFluor® 560 succinimidyl ester5605711200000.570.069100 ug71040
iFluor® 560 succinimidyl ester5605711200000.570.0695 mg71518
iFluor® 560 succinimidyl ester5605711200000.570.06910 mg71568
iFluor® 670 succinimidyl ester6716822000000.550.0331 mg1033
iFluor® 670 succinimidyl ester6716822000000.550.033100 ug71033
iFluor® 670 succinimidyl ester6716822000000.550.0335 mg71512
iFluor® 670 succinimidyl ester6716822000000.550.03310 mg71562
iFluor® 460 succinimidyl ester46849380000∼0.80.461 mg1022
iFluor® 460 succinimidyl ester46849380000∼0.80.46100 ug71022
iFluor® 460 succinimidyl ester46849380000∼0.80.465 mg71502
iFluor® 460 succinimidyl ester46849380000∼0.80.4610 mg71552
iFluor® 440 succinimidyl ester434480400000.670.2291 mg1041
iFluor® 440 succinimidyl ester434480400000.670.229100 ug71041
iFluor® 440 succinimidyl ester434480400000.670.2295 mg71519
iFluor® 440 succinimidyl ester434480400000.670.22910 mg71569
iFluor® 665 succinimidyl ester667692110,0000.220.09100 ug1550
iFluor® 665 succinimidyl ester667692110,0000.220.091 mg1551
iFluor® 665 succinimidyl ester667692110,0000.220.095 mg1552
iFluor® 665 succinimidyl ester667692110,0000.220.0910 mg1553
iFluor® 690 succinimidyl ester6857042200000.30.061 mg1042
iFluor® 720 succinimidyl ester7167402400000.140.131 mg1046
iFluor® 740 succinimidyl ester7427642250000.20.161 mg1047
iFluor® 597 succinimidyl ester5986181000000.70.5141 mg1050
iFluor® 770 succinimidyl ester7777972500000.160.081 mg1369
iFluor® 780 succinimidyl ester7848082500000.160.121 mg1371
iFluor® 830 succinimidyl ester830867---1 mg1384
iFluor® 570 succinimidyl ester5605711200000.580.0691 mg1034
iFluor® Ultra 594 succinimidyl ester5866011800000.910.051 mg71650
iFluor® Ultra 594 succinimidyl ester5866011800000.910.05100 ug71651
iFluor® Ultra 594 succinimidyl ester5866011800000.910.055 mg71652
iFluor® Ultra 647 succinimidyl ester6556702500000.390.071 mg71670
iFluor® Ultra 647 succinimidyl ester6556702500000.390.07100 ug71671
iFluor® Ultra 647 succinimidyl ester6556702500000.390.075 mg71672
iFluor® Ultra 750 succinimidyl ester7497732500000.320.051 mg71680
iFluor® Ultra 750 succinimidyl ester7497732500000.320.05100 ug71681
iFluor® Ultra 750 succinimidyl ester7497732500000.320.055 mg71682
ReadiCleave™ SSL biotin NHS ester5 mg3030
ReadiCleave™ AML Cy5 NHS ester6516702500000.031 mg7000
ReadiCleave™ XFD532 AML-NHS ester534553810000.611 mg7001
ReadiCleave™ ROXtra AML-NHS ester1 mg7002
ReadiCleave™ iFluor 700 AML-NHS ester6907132200000.230.041 mg7003
ReadiCleave™ FITC AML-NHS ester498517800000.790.2751 mg7004
ReadiCleave™ iFluor 488 AML-NHS ester491516750000.90.111 mg7005
ReadiCleave™ iFluor 546 AML-NHS ester5415571000000.670.151 mg7006
ReadiCleave™ iFluor 594 AML-NHS ester5886041800000.530.041 mg7007
ReadiCleave™ XFD488 AML-NHS ester499520730000.920.111 mg7008
ReadiCleave™ XFD546 AML-NHS ester5615721120000.790.121 mg7009
ReadiCleave™ XFD647 AML-NHS ester6506712390000.330.031 mg7012
ReadiCleave™ iFluor 488 SSL-NHS ester491516750000.90.111 mg7040
ReadiCleave™ FAM SSL-NHS ester491516730000.920.351 mg7041
ReadiCleave™ Cy5-SSL-NHS ester6516702500000.270.031 mg7050
ReadiCleave™ XFD532 SSL-NHS ester534553810000.610.091 mg7051
ReadiCleave™ ROXtra SSL-NHS ester1 mg7052
ReadiCleave™ iFluor 700 SSL-NHS ester1 mg7053
AzoDye-3 Succinimidyl Ester5 mg2472
mFluor™ Violet 530 SE393543200001 mg1616
AzoDye-1 PEG10 Succinimidyl Ester--5 mg2403
IDA succinimidyl ester--5 mg12632
NTA succinimidyl ester--5 mg12608
RatioWorks™ PH165, NHS ester463641--1 mg21214
RuFluor™ succinimidyl ester----1 mg1520
3-Maleimidopropionic acid N-hydroxysuccinimide ester *CAS 55750-62-4*----100 mg4507
  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. φ = fluorescence quantum yield.
  3. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).

Sulfonyl Chlorides

Sulfonyl chloride labeling mechanism
Figure 2. The reaction of a sulfonyl chloride compound with a primary amine.
Sulfonyl chlorides are highly reactive compounds for amine modification. Reaction of a sulfonyl chloride reagent with a protein or other amine-containing molecule results in the loss of the chlorine atom and the formation of a very stable sulfonamide bond. Although the stability of sulfonamide bonds are strong enough to withstand complete protein hydrolysis, sulfonyl chlorides are much more challenging to use and, therefore, not recommended for routine protein labeling. Conjugation occurs optimally under alkaline conditions (pH 9-10) in phosphate, bicarbonate, and borate buffers. Sulfonyl chlorides are quite unstable in aqueous solutions, especially at higher pH levels, and should never be used with the solvent dimethylsulfoxide (DMSO), as it reacts with them.

Table 2. Fluorescent dye sulfonyl chlorides for labeling proteins, antibodies and peptides.

Ex (nm)
Em (nm)
CF at 280³
Unit Size
Cat No.
Lissamine Rhodamine B Sulfonyl Chloride [Sulforhodamine B sulfonyl chloride] *CAS 62796-29-6*558575850000.330.171100 mg470
Sulforhodamine 101 sulfonyl chloride [Texas Red®] *CAS 82354-19-6*5866031160000.930.3610 mg480
Dansyl chloride [5-Dimethylaminonaphthalene-1-sulfonyl chloride] *CAS 605-65-2*33551833000.700.387100 mg811
  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. φ = fluorescence quantum yield.
  3. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).


Isothiocyanate labeling mechanism
Figure 3. The reaction of an isothiocyanate compound with a primary amine.
Isothiocyanates are moderately reactive electrophilic compounds that react with amino, hydroxyl, or thiol groups to form thioureas, O-thiocarbamates, or dithiocarbamates. In the reaction, the central, electrophilic carbon of the isothiocyanate moiety is attacked by an amine nucleophile. The resulting electron shift and proton loss create a thiourea bond between the isothiocyanate reagent and amine-containing compound with no leaving group. For isothiocyanate compounds, conjugation occurs optimally at alkaline pH levels (pH > 9), where the target amino groups are unprotonated (aromatic amines). Compared to bioconjugates modified with succinimidyl esters or sulfonyl chlorides, thiourea products are less stable and are more susceptible to deterioration over time. Popular isothiocyanate derivatives include fluorescein isothiocyanate (FITC) and tetramethylrhodamine isothiocyanate (TRITC) for labeling proteins, particularly antibodies, with green or orange/red fluorescence, respectively.

Table 3. Fluorescent dye isothiocyanates for labeling proteins, antibodies, and peptides.

Ex (nm)
Em (nm)
CF at 280 nm³
Unit Size
Cat No.
5-FITC [Fluorescein-5-isothiocyanate] *CAS 3326-32-7*491516730000.920.254100 mg120
5-FITC [Fluorescein-5-isothiocyanate] *CAS 3326-32-7*491516730000.920.2541 g121
5-FITC [Fluorescein-5-isothiocyanate] *CAS 3326-32-7*491516730000.920.25410 g122
5(6)-TRITC [Tetramethylrhodamine-5-(and-6)-isothiocyanate] *CAS 95197-95-8*5445701000000.100.3425 mg409
5(6)-TRITC [Tetramethylrhodamine-5-(and-6)-isothiocyanate] *CAS 95197-95-8*5445701000000.100.345 mg410
5-TRITC [Tetramethylrhodamine-5-isothiocyanate] *CAS 80724-19-2*5445701000000.100.341 mg415
5-TRITC [Tetramethylrhodamine-5-isothiocyanate] *CAS 80724-19-2*5445701000000.100.345 mg416
6-TRITC [Tetramethylrhodamine-6-isothiocyanate] *CAS 80724-20-5*5445701000000.100.341 mg417
6-TRITC [Tetramethylrhodamine-6-isothiocyanate] *CAS 80724-20-5*5445701000000.100.345 mg418
  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. φ = fluorescence quantum yield.
  3. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).


Optimal Bioconjugation with Amine-Reactive Fluorophores

Qualities of a desirable bioconjugate include one that is highly fluorescent, less susceptible to photobleaching and exhibits good water solubility while retaining the same functional properties as its innate unmodified counterpart. Two factors that contribute considerably to achieving these qualities include the fluorophore itself and the number of dyes attached to the protein, more commonly referred to as the degree of labeling (DOL).

Fluorophores with high extinction coefficients and high quantum yields will generate bright fluorescence signals upon excitation, and the higher these values are, the brighter the fluorophore. For instance, phycoerythrin (PE), which has an extinction coefficient of 1,960,000 cm-1M-1, is one of the brightest fluorescent dyes known and is widely used to image low-abundance cellular targets in flow cytometry. However, due to its rapid photobleaching, PE has limited use in immunofluorescence microscopy. FITC, another widely used labeling dye for biological research, benefits from high absorptivity, excellent fluorescent quantum yield, and good water solubility, but its high rate of photobleaching and pH-sensitive fluorescence can affect its performance, especially for staining protocols using acidic buffers. New generation dyes, such as iFluor® dyes and Alexa Fluor® dyes, are highly photostable and maintain high fluorescence intensities over a broad pH range. The enhanced photostability and high quantum yield of iFluor® 488 (φ=0.9) enable the detection of biological targets with greater sensitivity.

More important than selecting the right fluorophore for any experiment is optimizing the degree of labeling such that the bioconjugate retains its functionality and has a high fluorescence output. Since proteins contain numerous primary targets for amine modification, the amount of dye-labeled to a single protein can vary significantly. Conjugates with a low DOL may result in weak fluorescence, while a high DOL can cause conjugates to precipitate out of solution, bind nonspecifically, lose functionality, and self-quench. For antibodies, the optimal DOL usually ranges between 2 to 10. A more precise value, however, will largely depend on the properties of the label and protein. This means that for many bioconjugations, the optimal DOL must be experimentally determined, often through several small-batch labelings. For accurate determination of dye:protein molar ratios, use AAT Bioquest's DOL calculator

After conjugation, removing as much reactive-free dye as possible is essential. This can be done via gel filtration chromatography, dialysis, or using a desalting column, such as the ReadiUse™ Disposable PD-10 desalting column.

Fluorescence Spectrum Viewer

Need assistance selecting the best fluorophore for your experiment, use our Fluorescence Spectrum Viewer:
  • View and compare fluorophores and fluorescent proteins for biological applications
  • Check spectral compatibility
  • Add multiple excitation and emission filters
  • Save spectra configuration as a PNG or hyperlink