Getting started with RAMP

How do you get started with RAMP?  Recently I sent an e-mail to a researcher with this question on bycatch in a purse seine.  My answer follows.  Good to hear from you.  Sounds like you have some bigger fish to work with.  Sample sizes depend on what you are going to accomplish and the effect size anticipated.  1) The first tasks are to get fish in good condition, lean how to hold them in good condition, and then to establish a RAMP curve.  Get as many fish as you can comfortably hold and maintain.  Then 2) establish reflex actions that can be tested consistently and that respond consistently and strongly.  Find as many testable reflex actions as you can as this makes a more refined RAMP score.  See Davis 2010 and Raby et al. 2012 papers for detailed information.  For larger salmon that are difficult to handle, Raby tested without a restraining device: 
"Each reflex was assessed categorically (0 = unimpaired, 1 = impaired) in a conservative matter – that is, if the handler had doubt as to whether the reflex was present, it was recorded as being impaired. Reflexes tested were the following: tail grab, body flex, head complex, vestibular-ocular response (VOR) and orientation. Presence of the tail grab response was assessed by the handler attempting to grab the tail of the fish with the fish submerged in water (in a fish bag or holding trough); a positive response was characterized by the fish attempting to burst-swim immediately upon contact. The body flex response was tested by holding the fish out of water using two hands wrapped around the middle of the body. The fish actively attempting to struggle free was characterized as a positive response. Head complex was noted as positive if, when held out of water, the fish exhibited a regular pattern of ventilation (for 5 s) observable by watching the opening and closing of the lower jaw. VOR was observed by turning the fish on its side (i.e. on a lengthwise axis) out of water. Positive VOR was characterized by the fish’s eye rolling to maintain level pitch, tracking the handler. Finally, upon release, each fish was placed upside-down in the river just below the surface: a positive orientation reflex was noted if the fish righted itself within 3 s. The entire reflex assessment took 20 s to complete and was always conducted on fish upon release. If a fish was too vigorous to allow researcher handling and assessment of reflexes, it was assigned an unimpaired status for all reflexes."  
Then 3) once consistent reflexes are identified and tests are finalized, a RAMP curve is constructed which establishes the relationship between reflex impairment and mortality.  This is the scoping process that establishes what stressor intensities will kill fish and what are sub-lethal and produce lesser RAMP scores.  This process involves exposing fish to a range of capture stressor intensities of interest.  For purse seine capture this may include net abrasion time (extent of injury) and air exposure time (extent of hypoxia) and temperature if there are enough fish to test.  The stressor range should induce reflex impairment and mortality that ranges from no impairment and mortality to complete impairment and mortality.  In this way, the RAMP curve covers all possible outcomes for the fish.  If possible, the most efficient use of limited fish is to expose and tag individual fish so that for each individual, a RAMP score can be assigned to a specific stressor intensity.  4) Once you have a RAMP curve, you can test fish in the field for RAMP, note their capture conditions, and predict mortality based on RAMP and also on capture conditions.  

Initially you will need at least 10 fish to construct a RAMP curve, more (20 fish) is better as it will give you a curve with tighter confidence bounds.  For a simple example lets say that fish die after 15 min in air.  So exposures might look like this: fish 1- 0 min air, fish 2 - 2 min air, fish 3 - 4 min air, fish 4 - 6 min air, fish 5 - 8 min air, fish 6 - 10 min air, fish 7 - 12 min air, fish 8 - 14 min air, fish 9 - 16 min air, fish 10 - 18 min air.  If injury is important you might do a combination of net holding and air exposure.  Expose the fish to the assigned stressor, test the fish for reflex impairment, then hold the fish and observe for delayed mortality.  Holding time vary with species and usually range from 3-11 days.