Optical tweezers that not always atract but also repel

[Fioda]

Hi!
I have the following problem: the optical tweezers attract beads but when the beads get to the trap, they are repelled. Furthermore, when once in a while a bead gets trapped, it goes out of focus.
The tweezers are coupled to an inverted optical microscope. They are made with an IR laser and an aquous immersion 100X objective (NA 1)
Some trick of what to do?

 

[Andy Resnick]

My first guess (based on the information you provide) is that the trap is attractive in 2 dimensions (in the plane of focus) but not in 'z'. Generally, this is caused by poor alignment or incomplete control of residual aberrations, typically spherical and astigmatism.

Directly imaging the trap is essential to ensure good alignment- place a mirror at the sample plane and as you scan through focus, you will easily be able to tell if you have alignment problems.

Edit- your statement that trapped beads appear 'out of focus' also indicate that your tweezer beam isn't properly matched to the objective lens- either not collimated (for infinity-corrected objectives) or otherwise incorrectly aligned.

 

[Fioda]

Hi Andy, thanks for your answer. I did what you told me (using a viewer card). I finally could trap beads again BUT out of focus... besides, the laser does not look perfect (I attached a photo). How can I improve the focus of the laser? How can I get rid of residual aberrations?

 

[Andy Resnick]

yikes... I assume this is the image of the focal spot obtained using a first-surface mirror?

I can see astigmatism and coma- if the spot moves from side to side as you pass through focus, that's confirmation of off-axis aberrations. Those aberrations are often caused by angular misalignment- I have to align the tweezer to within a few arcminutes, and sub-arcmin is ideal.

Ideally, you should see an Airy disc on focus, and concentric circles off-focus- I should be able to post a few pics shortly.

Edit- here they are: one near focus, one past focus
http://imageshack.us/a/img41/7821/focus0000.jpg
http://imageshack.us/a/img715/741/pastfocus0001.jpg

The vertical stripes (and satellite dots) are, I think, from the dichroic mirror used to couple the tweezer to the microscope. It does not appear to degrade trap performance.

 

[Fioda]

Thanks again!
Here are the images of the airy discs. They are not so bad, are they?
However, the trap is still not working properly, and when the beads are trapped (most of the time they go out of the trap but sometime they stay there), they are out of focus. Probably, I can overcome that problem moving the collimation of the beam expander?