What am I doing wrong?

Also, although it may not be the problem in this specific instance, depending how many tubes and the focal length, you are going to want to stop down because DOF gets really short when working in high manification.
 
If shooting from mirror lock up diesn't work, try a couple of other things.

It seems that the lower right hand quarter (no pun intended) is pretty sharp, but the rest of the image is OOF. With macro shots the DOF is teeny tiny in lots of cases. Are you sure the camera is perfectly square to the subject? That can have a big effect.

Next, I think the D5000 has live view, right? I find it much easier to focus through live view in macro situations than I do through the viewfinder. You might want to give that a try too.
 
I've the got the sharpness I want/need through the viewfinder though. It's only after I take the picture that it looks like that.
There is a filter in front of the image sensor. It is known as a low-pass or anti-aliasing (AA) filter.

When you look in the viewfinder, you are not seeing what the photo will look like when the mirror moves up out of the light path so the light can go through the AA filter to the image sensor.

The AA filter softens the focus, and images need to be sharpened in post processing to counter the AA effects.

Anti-aliasing filter - Wikipedia, the free encyclopedia
 
This may sound odd, but try a different shutter speed... longer or shorter.

I noticed years ago that this could happen when I was shooting around 1/15 sec.
It SEEMS it's camera shake. Shooting at 1 sec did better.

I can only guess it was because the mirror was moving for a smaller % of the total exposure.

I dunno. Maybe it's all in my head. But try it and let me.

-Pete

DING DING DING!!! 1/15s is right in the range of shutter speeds that are most affected by mirror slap, for the reason mentioned above. I had this exact question a while back, and got some great advice. Here's the thread:http://www.thephotoforum.com/forum/beyond-basics/217745-long-exposures-motion-blur-tripod-vibration.html
 
Well, I answered my own question via google and set the exposure display to 2 seconds and I feel like I came up with much better results than the first photo.

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Thanks for all your help guys.
 
Well, unfortunately the D5000 doesn't have the capability to lock-up the mirror. Would using the exposure delay effectively do the same task?

This will help reduce camera and tripod vibration and it's good practice for tripod shots in general, but will do nothing to reduce mirror slap, as the mirror still flips up and back down at the same time it normally would. Incidentally, I have the same limitation on my D40, and it's really just a silly, deliberate firmware omission for the cheaper cameras. You could always try doing a much longer exposure with F/11-F/16; this may help you two-fold: 1. smaller aperture yields greater depth of field and overall sharper lens performance (don't go too high with the F number...diffraction can reduce sharpness beyond F/16 or so); 2. slower shutter speed mitigates effect of mirror slap. Of course, if you're using a flimsy tripod, the long exposure could be counter-productive. Use a remote to trigger the exposure, or alternatively use a 10s delay. DO NOT MOVE during the exposure. Foot falls, or bumping the table with your setup can create substantial vibration which will blur the shot.
 

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