From a 3.3v Pin do we separate Positive and Negative flow? If so, how?
The 3.3V connection on a RPi is +3.3 volts, relative to the Pi's Ground.  The Pi also has a +5V connection on the I/O header - this is +5V relative to the Pi's ground pins.

In general, any voltage you see mentioned on a circuit is the voltage relative to the circuit's ground.  And, in most cases, the term "ground" does not imply a connection to the earth - it is just the point in the circuit that we choose to call "Zero Volts", and use as a reference when measuring voltages elsewhere in the circuit.