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Fishery Bulletin 94(4), 1 996 



lar weight standards (Sigma Chemical) are described 

 in Schultz and Clarke (1995). 



Glycoproteins were identified on electroblots by 

 using the method of O'Shannessy et al. (1987). 



Results 



The steps for purification of a 66 kDa protein from a 

 soluble saline extract of L. griseus are shown in Table 



1. Figure 1 depicts samples of each purification step 

 in SDS-PAGE (gel A) and corresponding Western 

 blots (gels B and C) leading to the highly purified 

 protein. The protein was a trace component of the 

 saline extract ( arrow, Al ) but was more concentrated 

 with successive purification steps (A, 2-4). The steps 

 of purification included L. griseus saline extract in 

 lane 1, the A.S. (0.38 gm/mL extract) supernatant 

 solution of the extract in lane 2, the 0.27 M NaCl 

 eluate from a Mono S cation exchange column after 



KDa 



12 3 4 1 2 3 4 



It 



A B C 



Figure 1 



Samples from the purification steps of soluble L. griseus extract leading to the highly 

 purified 66 kDa protein in lane 4. Gel (A) shows samples after SDS-PAGE stained with 

 Coomassie brilliant blue. The corresponding Western blots were developed by using 

 biotinylated goat anti-whole L. griseus antiserum (B) and goat anti-L. griseus 66 kDa 

 protein IgG (C). Reference standard proteins ikDa) are shown in the stained gel. The 

 arrows in gels A and C show the position of the 66 kDa protein. Numbered lanes denote 

 the following: 1) whole L. griseus extract (235 ug total protein); 2) ammonium sulfate (0.38 

 g/mL extract) supernatant solution of L. griseus whole extract (228 ug); 3) eluate from 

 Mono S cation exchange column (20 ug) after FPLC; and 4) eluate from Mono Q anion 

 exchange column (0.32 ug) after FPLC. Note that the reaction of biotinylated goat anti- 

 whole L. griseus antiserum with the 66 kDa L. griseus protein was not detected in B. 



