Dear Kumar,

the attached link may help you to find a less harsh and more effective route for the first step of your synthesis. Unfortunately the second step the reaction described within the mentioned contribution is more than unlikely in this way (think about Sol-Gel-synthesis). APTES is a highly reactive silane which prefers to form mono-aminopropyl siloxanes especially if water and acid are present. A reason for the adhesion may be the change is surface tension of the PS by forming Nitro-PS and the surplus of acid still remaining attached to/into the polymer due to the fabrication. Why do you not reduce the Nitro-functionality to an amine?

http://www.sciencemadness.org/talk/files.php?pid=127922&aid=5534

Dear Sir Schmitt,

thanks for the suggestion sir, Actually i am not using APTES rather i have 3-marcapto  propyltrimethoxy silane i.e MPTS. i want to prepare a gold/silver nanoparticle layer over MPTS SAM. i am facing 2 major problems

1. polystyrene cuvette loosing transparency after nitration (though i will try your suggestion as given in the paper you have reffered and will reply soon with the results) 

2. the second problem is that there is not a single protocol available for MPTS SAM over PS surface. so i am confused wether i go for a ethanol:water (95:5)  solution of MPTS(10%) or go for a sol-gel kind of MPTS treatment over PS surface method. or should i try this out with benzene MPTS solution(as prescribed by many authors in literature)

I further suggest using a glass cuvette (inorganic surface containing the necessary hydroxyl groups). Attached you will find the link to Gelest. The “Silicon compounds: silanes and silicones” catalogue will help you to clean your surface to apply silanes and to coat surfaces (section “silane coupling agents”).  

http://www.gelest.com/gelest/forms/GeneralPages/literature.aspx

Dear Sir Schmitt

Thank you for your valuable suggestions and time.

I will try to implement your suggestion and let you know about the results. 

Thanks again

Alkoxysilanes will condense with surface hydroxyl groups and other silanols to form siloxane bonds, especially in the presence of mM acetic acid or other catalyst.  A plasma treatment of your polystyrene surface should produce such OH groups.  I don't know which plasma will work best nor the treatment time and pressure, but I would try an oxygen plasma or a water vapor plasma. Simply treat the polystyrene plates in a small plasma cleaner charged with the appropriate atmosphere.  XPS analysis of the surface afterwards would reveal the presence of OH groups.

Thanks Walter Sir for Suggestion. Actually I am looking for a chemical reagent based protocol of -OH group generation as well. if you can suggest any good protocol for generating -OH group over polystyrene that would be much helpful.

thanks in advance

Just out of curiosity, did all of the well plates come from the same lot?  I once had an optimized procedure to silanize "glass microscope slides" from a major supplier.  Then one day the procedure wasn't optimized.  Eventually, I found out that the supplier had switched manufacturers and there was something different about the surface chemistry of the slides.  If I recall correctly, there are a range of PS well plates available, including some with surface modifications for ELISA.

Coolbaugh Sir, 

No Sir, I haven't change the material. i used ELISA plates from the same slot. 

http://www.lifescienceglobal.com/pms/index.php/jbas/article/viewFile/2481/1465

You may be insterested in the paper in the link above.  I have modified PS surfaces using a 30s treatment with a chromic acid/sulfuric acid mix.  This creates a surface with a negative charge at high pH.  You could then treat with poly(allylamine hydrochloride) (PAH) which would be suitable for Au nanoparticle immobilization.