Systems, apparatuses and methods for simultaneous multi-plane imaging

A method, comprising:

• generating an optical signal including a set of pulses at a first repetition rate, wherein the first repetition rate is an n-multiple of a second repetition rate, wherein the set of pulses at the second repetition rate include n sets of pulses, each set of pulses of then set of pulses having a different polarization state than each other of the n set of pulses and operating at the first repetition rate;
• splitting the optical signal into a set of n excitation signals to excite a sample, each excitation signal of the set of n excitation signals corresponding to a set of pulses of the n set of pulses;
• focusing each excitation signal of the set of n excitation signals to a different focal plane of the sample than each other excitation signal of the set of n excitation signals to generate a response signal in response to the set of n excitation signals;
• and
• separating the emission from the different focal planes in the response signal into a set of n emission signals, each emission signal of the set of n emission signals corresponding to an excitation signal of the set of n excitation signals.

A system, comprising:

• a laser source to generate an optical signal, the optical signal including a set of pulses at a first repetition rate;
• an electro-optic modulator and a polarizing beam splitter to receive the optical signal and to generate a multiplexed optical signal including a set of pulses at a second repetition rate, wherein the second repetition rate is twice the first repetition rate, wherein the set of pulses at the second repetition rate include a first set of pulses and a second set of pulses each at the first repetition rate, the first set of pulses having a different polarization state than the second set of pulses;
• a focusing unit to receive the multiplexed optical signal and to split the multiplexed optical signal into a first excitation signal and a second excitation signal to excite a sample, the first excitation signal corresponding to the first set of pulses, the second excitation signal corresponding to the second set of pulses, the focusing unit including a first set of focusing optics and a second set of focusing optics having a different longitudinal axis than the first set of focusing optics, each of the first set of focusing optics and the second set of focusing optics including:
• a quarter wave plate;
• a second objective;
• and
• a focusing mirror to independently adjust a position of the focal plane of its corresponding excitation signal;
• an objective to receive the first excitation signal and a second excitation signal and optically coupled to the sample to illuminate the sample and to generate an emission from the sample, wherein the first excitation signal is focused on a first focal plane of the sample and the second excitation signal is focused on a second focal plane of the sample that is different than the first focal plane;
• a detector configured to generate a response signal based on the emission from the sample;
• and
• a switch to separate the response signal into a first emission signal corresponding to the first excitation signal and a second emission signal corresponding to the second excitation signal.