b'Alexa Fluor and Alexa Fluor Plus secondary antibodies The wide range of Invitrogen Alexa Fluor and Alexa Fluor Plus secondary antibodies for western blottingA 130 kDaprovides multiple options of nonoverlapping spectra95 kDa Hsp90to better facilitate multiplex analysis. Multiple targets70 kDa HDAC1can be detected independently in the same lane and55 kDablot with clearly distinguishable colors. Alexa Fluor Plus43 kDasecondary antibodies combine enhanced sensitivity and 5.0 g 1.6 g 0.6 g 0.2 g15.0 g 0.06 glow background for higher signal-to-noise ratios and better multiplexing results.B 130 kDaFeatures: 95 kDa Hsp90 Detection of low-abundance targets; up to 5.8 times70 kDa HDAC1higher signal-to-noise ratio in fluorescent western blotting 55 kDa43 kDa High cross-adsorption to minimize cross-reactivity for low Figure 39. Comparison of detection sensitivity. (A) Invitrogen Alexa background and better multiplexing results Fluor Plus 800 Secondary Antibody shows more bands at higher dilutions, demonstrating greater sensitivity and range of detection compared to (B)Improved sensitivity and range of linear detection toanother suppliers fluorophore-conjugated secondary antibody.provide more detail (Figure 39) Increased photostability and superior brightnessAlexa Fluor Plus secondary antibodies are available for key target species and are labeled with a wide range of superior dyes across the color spectrum (488800 nm) for multiplex western blotting applications (Table 23).Table 23. Alexa Fluor Plus secondary antibodies are available in a wide range of target species and wavelengths.Alexa FluorAlexa FluorAlexa FluorAlexa FluorAlexa FluorAlexa FluorAlexa Fluor Plus 405 Plus 488 Plus 555 Plus 594 Plus 647 Plus 680 Plus 800Goat anti-rabbit A48254 A32731 A32732 A32740 A32733 A32734 A32735Goat anti-mouse A48255 A32723 A32727 A32742 A32728 A32729 A32730Goat anti-chicken A48260 A32931 A32932 A32759 A32933 A32934 A32935Goat anti-rat A48261 A48262 A48263 A48264 A48265Goat anti-human A48275 A48276 A48277 A48278 A48279Donkey anti-goat A48259 A32814 A32816 A32758 A32849 A32860 A32930Donkey anti-rabbit A48258 A32790 A32794 A32754 A32795 A32802 A32808Donkey anti-mouse A48257 A32766 A32773 A32744 A32787 A32788 A32789Did you knowAll Alexa Fluor Plus secondary antibodies are cross-adsorbed against potential cross-reactive species. Cross-adsorption or pre-adsorption is a purification step to increase specificity of the antibody resulting in less background staining and cross-reactivity. The secondary antibody solution is passed through a column matrix containing immobilized serum proteins from potentially cross-reactive species. Only the nonspecific-binding secondary antibodies are captured in the column, and the highly specific secondary antibodies flow through. Further passages through additional columns result in highly cross-adsorbed preparations of secondary antibody. The benefits of these extra steps are apparent in multicolor-staining experiments where there is potential cross-reactivity with other primary antibodies.61'